Bacillus subtilis

J Bacteriol, 1979 May, 138(2), 370 - 6
Characterization of a succinate dehydrogenase complex solubilized from the cytoplasmic membrane of Bacillus subtilis with the nonionic detergent Triton X-100; Hederstedt L et al.; A succinic dehydrogenase (SDH) complex has been purified from Triton X-100-solubilized membranes from Bacillus subtilis by precipitation with specific antibody . Radioactively labeled precipitated complex was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by autoradiography of the gels . The complex contained equimolar amounts of three polypeptides with approximate molecular weights of 65,000, 28,000, and 19,000 . Five succinic dehydrogenase-negative mutants, belonging to the citF group, contained the 65,000-dalton polypeptide in a soluble form in the cytoplasm . Each 65,000-dalton polypeptide had about one molecule of flavin bound . Another citF mutant, citF11, which lacks the 65,000-dalton polypeptide, contained a membrane-bound 28,000-dalton polypeptide . The wild-type succinic dehydrogenase complex contained cytochrome, probably a cytochrome b . The 19,000-dalton polypeptide is suggested to represent the apoprotein of this cytochrome . The 65,000-dalton and the 28,000-dalton polypeptides are thought to constitute succinic dehydrogenase and to correspond to the flavoprotein and the ironprotein, respectively, as described for succinic dehydrogenase isolated from beef heart mitochondria or Rhodospirillum rubrum chromatophores . The results presented suggest that in B . subtilis succinic dehydrogenase is attached to a cytochrome b in the membrane via the 28,000-dalton (ironprotein) polypeptide.

J Bacteriol, 1979 May, 138(2), 345 - 51
Export of extracellular levansucrase by Bacillus subtilis: inhibition by cerulenin and quinacrine; Caulfield MP et al.; Bacillus subtilis B secretes an inducible, extracellular enzyme, levansucrase . Inhibition studies were undertaken to investigate the possible mechanism of release of this enzyme . The antibiotic cerulenin, at a concentration of 10 micrograms/ml, totally inhibited de novo lipid synthesis in B . subtilis B for at least 1 h, while only slightly reducing protein and RNA synthesis . At this concentration cerulenin, added concomitantly with the inducer sucrose, prevented the release of levansucrase for at least 150 min . This was not due to the prevention of inducer uptake by the cells . The release of the enzyme was also independent of cell division . In B . subtilis 1007 the induction of beta-galactosidase by 5 mM lactose was not prevented by cerulenin . Preliminary evidence indicated the association of a lipid moiety with the enzyme as it passes through the cytoplasmic membrane . Quinacrine (0.2 mM), which inhibits the penicillinase-releasing protease of Bacillus licheniformis, inhibited levansucrase release from B . subtilis B, but had no effect on lipid synthesis.

J Bacteriol, 1979 May, 138(2), 314 - 9
Specific inhibition of outgrowth of Bacillus subtilis spores by novobiocin; Gottfried M et al.; Spores of a Bacillus subtilis mutant temperature sensitive in deoxyribonucleic acid (DNA) replication proceeded through outgrowth at the nonpermissive temperature to the same extent as the wild-type parent spores . In contrast, the DNA synthesis inhibitor novobiocin completely prevented spore outgrowth while displaying a marginal effect on logarithmic growth during one generation time . Inhibition of outgrowth by novobiocin occurred in the absence of DNA replication, as demonstrated in an experiment with spores of the temperature-sensitive DNA synthesis mutant at the restrictive temperature . Novobiocin inhibited the initial rate of ribonucleic acid synthesis to the same extent in germinated spores and in exponentially growing cells . A novobiocin-resistant mutant underwent normal outgrowth in the presence of novobiocin . Therefore, novobiocin inhibition was independent of its effect on chromosome replication per se.

Biochim Biophys Acta, 1979 Apr 19, 552(3), 558 - 62
Interactions between bacterial membranes and peptidolipids: lysis of micrococcus luteus protoplasts by derivatives of peptidolipidic antibiotics from bacillus subtilis; Besson F et al.; The lysis of protoplasts of Micrococcus luteus has been tested with various derivatives of three peptidolipidic antibiotics: iturin A, mycosubtilin and bacillomycin L . The lytic activity is dependent to the nature of the substituting group and to the position of the substituted aminoacid residue . The acetylation of OH groups leads to a decrease of the lytic activity of the natural antibiotics . The methylation of aspartyl residues of bacillomycin L gives a strong lytic activity while natural bacillomycin L has no lytic activity . The methylation of the tyrosyl residue enhances the lytic activities of iturin A and of bacillomycin L-dimethyl ester and reduces that of mycosubtilin . Correlations between the structures of derivatives and their lytic action on M . luteus protoplasts are discussed.

Biochim Biophys Acta, 1979 Apr 19, 552(3), 492 - 8
Characterization of glutamate transport system in hydrophobic protein (H protein) of Bacillus subtilis; Kusaka I et al.; Hydrophobic protein (H protein) was isolated from membrane fractions of Bacillus subtilis and constituted into artificial membrane vesicles with lipid of B . substilis . Glutamate was accumulated into the vesicle when a Na+ gradient across the membrane was imposed . The maximum effect of Na+ on the transport was achieved at a concentration of about 40 mM, while the apparent Km for Na+ was approximately 8 mM . On the other hand, Km for glutamate in the presence of 50 mM Na+ was about 8 micro M . Increasing the concentration of Na+ resulted in a decrease in Km for glutamate, maximum velocity was not affected . The transport was sensitive to monensin (Na+ ionophore) . Glutamate was also accumulated when pH gradient (interior alkaline) across the membrane was imposed or a membrane potential was induced with K+-diffusion potential . The pH gradient-driven glutamate transport was sensitive to carbonylcyanide m-chlorophenylhydrazone and the apparent Km for glutamate was approximately 25 microM . These results indicate that two kinds of glutamate transport system were present in H protein: one is Na+ dependent and the other is H+ dependent.

Experientia, 1979 Apr 15, 35(4), 470 - 2
Different effects of D-glucose anomers for respiration of bacterial germinated spores; Ishihara H et al.; Effects of alpha- or beta-D-glucose on the respiration of germinated spores (only germinated spores not including swollen spores and elongated spores) of Bacillus subtilis and B . megaterium were studied . In our conditions, net amount of oxygen consumed by 10(10) germinated spores of B . subtilis per min after addition of alpha- or beta-D-glucose was 1.6 microgram or 6.6 microgram (beta/alpha = 4.13), while that by B.megaterium was 4.5 microgram of 6.8 microgram (beta/alpha = 1.51), respectively . However, the net amounts of oxygen consumed by 10(10) vegetative cells per min after addition of alpha- or beta-D-glucose were identical, for B.subtillis in both cases 443.0 microgram and for B.megaterium in both cases 604.4 microgram.

J Biol Chem, 1979 Apr 10, 254(7), 2184 - 6
Regulation of teichoic acid synthesis during phosphate limitation; Glaser L et al.; Bacillus subtilis W-23, when placed in phosphate-free medium, ceases to synthesize teichoic acid and synthesizes teichuronic acid . The enzymatic basis for the cessation of teichoic acid synthesis is the irreversible inhibition of the first membrane-bound enzyme involved in teichoic acid synthesis which catalyzes the reaction Undecapenol-P + UDP-GlcNAc leads to undecaprenol-P-P-GlcNAc + UMP.

J Gen Microbiol, 1979 Apr, 111(2), 353 - 61
Intrastrand self-complementary sequences in Bacillus subtilis DNA; Galloway DA et al.; Intrastrand self-complementary sequences have been isolated from the DNA of Bacillus subtilis by hydroxyapatite (HA) chromatography following thermal renaturation of strands separated by chromatography on methylated albumin kieselguhr (MAK) . The instrastrand structures derived from the MAK H strand (HA HII) were biologically active showing transforming activity for a wide variety of markers, as well as hybridization to both pulse-labelled and ribosomal RNA . Removal of regions of single-strand DNA with S1 nuclease did not significantly alter the biological activity of the self-annealed molecules . The overall efficiency of transformation and hybridization of the intrastrand self-annealing DNA was low suggesting that many sequences in the population are neither active in transformation to prototrophy nor transcribed into RNA.

Mol Gen Genet, 1979 Mar 5, 170(3), 345 - 9
Lethal effect of protamine and histone on competent Bacillus subtilis cells . Inhibition of genetic transformation by protamine in sublethal concentration; Antohi S et al.; Under experimental conditions of genetic transformation, protamine and total histone were bactericidal for Bacillus subtilis cells . The abilities to cause lethality were very similar for both, either protamine or histone, with no antagonistic effects amongst these natural polycations . With both basic proteins acting simultaneously the enhancement was higher than a summation of the separate lethal effects . Sublethal concentration of protamine added at the beginning of transformation time, produced a strong inhibition of transforming efficiency . The same concentration added later than 10 min from the start of transformation had no inhibitory effect . These facts together with the absence of inhibition by simple pretreatment of DNA alone as well as the cell protection by protamine against lytic activity of lysozyme, suggest a protamine-cell surface interaction which impedes DNA uptake events.

J Bacteriol, 1979 Mar, 137(3), 1452 - 5
Spectinomycin dependence in Bacillus subtilis; Henkin TM et al.; Spectinomycin dependence in Bacillus subtilis involves two mutations, one conferring drug resistance and the other producing a requirement for spectinomycin for growth.

J Gen Microbiol, 1979 Mar, 111(1), 165 - 80
Genetics analysis of spore germination mutants of Bacillus subtilis 168: the correlation of phenotype with map location; Moir A et al.; The isolation and characterization of 29 new germination (Ger) mutants of Bacillus subtilis 168 is described . These were classified, along with previously described mutants, into seven groups according to map location . The mutations in 26 GerA mutants mapped between cysB and thr; detailed mapping of two of these has located them very close to citG . These mutants were deficient in germination in alanine, but responded to the germinative combination of asparagine, glucose, fructose and KCl . One GerB mutant mapped on the origin-proximal side of hisA; it was normal in germination in alanine, but deficient in termination in a mixture of asparagine, glucose, fructose and KCl . Two GerC mutants were linked to lys, but were separable from a temperature-sensitive growth deficiency mapping between lys and trp . The GerC mutants had a similar germination phenotype to the GerA mutants . Three GerD mutants did not germinate in either of the above germinants or in Penassay Broth . They were located on the side of ery distal to cysA . The GerE mutant, which did not germinate in any of the three germinants, was located very close to citF and possessed an altered spore coat . The two GerF mutants were defective in germination in all three germinants and mapped on the origin proximal-side of hisA, but much closer to his than did the GerB mutant . A phosphoglycerate kinase-negative mutant altered in germination mapped between cysB and hisA (GerG) . These mutants have established a minimum of seven locations important to germination, and will be useful in the development and appraisal of theories of spore germination.

Nucleic Acids Res, 1979 Mar, 6(3), 1203 - 19
Antibody to B . subtilis DNA polymerase III: use in enzyme purification and examination of homology among replication-specific DNA polymerases; Barnes MH et al.; Bacillus subtilis DNA polymerase III (pol III), an arylhydrazinopyrimidine-sensitive, replication-specific enzyme, was used to generate a non-precipitating rabbit antibody which specifically inhibited pol III activity in vitro . The antibody was used to examine structural relationships among several DNA polymerases, and it was linked covalently to agarose; the antibody:agarose was employed to develop a rapid, selective method of purification of catalytically active B . subtilis pol III.

Mikrobiologiia, 1979 Mar-Apr, 48(2), 249 - 55
{Effect of exogenous nucleodepolymerases on Bacillus subtilis multiplication}; Kupriianova FG et al.; The effect of exogenous nucleodepolymerases on cell growth was studied with Bacillus subtilis . Nucleodepolymerases of microbial and animal origin stimulated growth of the culture . The stimulating action of DNAases depended on the dose of the enzyme added to the growth medium and on the phase of the cultural growth . Acceleration of the cultural growth correlated with intensification of DNA synthesis in the bacterial cells.

J Bacteriol, 1979 Mar, 137(3), 1354 - 61
Electron microscopic study of Bacillus subtilis protoplast fusion; Frehel C et al.; When protoplasts derived from sporulating cells of Bacillus subtilis were fused by exposure to polyethylene glycol (PEG) and fixed immediately thereafter, protoplasts with two enclosed prespores could be seen by electron microscope . The number of fusion events was greatly increased, and multiply fused protoplasts appeared, when the PEG-treated suspension was diluted in hypertonic broth and reincubated before fixation . This post-PEG incubation effect is taken to indicate a fusion mechanism of two steps: a short, PEG-dependent step of membrane activation, followed by a slow, metabolism-requiring step completing fusion . When prespore-bearing protoplasts from two genetically different strains were mixed and fused, the extent of fusion could also be followed by counting clones of recombinant bacteria . Maximal from the start, their number (1% of each parent type protoplast present) was unaffected by post-PEG incubation . Fusion in this case is apparently completed after plating on the wall-regeneration medium . After optimal post-PEG incubation, the majority of the protoplasts were seen to participate in fusion, and the cytological fusion observed, corrected for wall-regeneration frequency, accounted quantitatively for the prototrophic bacteria eventually recovered . These results are in good agreement with those obtained independently by Sanchez-Rivas and Garro (J . Bacteriol . 137:1340--1345, 1979).

J Bacteriol, 1979 Mar, 137(3), 1346 - 53
Parameters governing bacterial regeneration and genetic recombination after fusion of Bacillus subtilis protoplasts; Gabor MH et al.; Bacterial protoplast fusion, induced by polyethylene glycol, has been made more regular and convenient by further specification and improvement of various steps in the previously used procedure . These have made it possible to obtain regularly 100% regeneration of Bacillus subtilis cells from protoplasts before treatment with polyethylene glycol and yields of 10 to 75% from polyethylene glycol-treated protoplasts . Genetic recombination frequencies do not increase correspondingly . Also, when regeneration is reduced by various experimental conditions, recombination does not decrease in proportion . It is concluded that regeneration of recombinant-forming cells is independently determined and not closely related to the average regeneration for the population . Kinetic studies with varying individual parental or total protoplast concentrations strongly indicate that protoplast collision and contact is not the limiting factor determining the number of genetic recombinants obtained . Recombination approximates a linear, rather than quadratic, function of the total or of the majority protoplast population present, from which it is concluded that fusion events are always adequate to produce substantially more potential recombinants than are registered . The strong effect of the majority/minority ratio upon the number of minority cells that become recombinant is independent of which parent is in excess . This shows in a direct and physiological way that both parents are equivalent partners in their genetic contributions.

J Bacteriol, 1979 Mar, 137(3), 1340 - 5
Bacterial fusion assayed by a prophage complementation test; Sanchez-Rivas C et al.; In previous studies of bacterial protoplast fusion, only the frequencies of cell wall regeneration and of bacterial recombination were determined . In this work the frequency of the heterozygous fusion products is measured by prophage complementation . Two multiply marked nonsuppressing strains of Bacillus subtilis, each lysogenic for a different Sus mutant of the phage phi 105, were induced by mitomycin C, protoplasted, fused, and, after dilution in hypertonic broth, incubated until plating with phi 105-sensitive indicator bacteria . When cell lysis was avoided, the frequency of the heterozygous fused cells could be determined from the number of infectious centers produced . The very high frequencies observed are in good agreement with those determined directly, with nonlysogenic strains, by electron microscopic examination of the fused protoplasts (C . Frehel, A . M . Lheritier, C . Sanchez-Rivas, and P . Schaeffer, J . Bacteriol . 137:1354--1361, 1979) . Evidence is presented that fusion occurs in two steps, one polyethylene glycol dependent, the other energy requiring . The bacterial growth medium affects the ability of the protoplasts to fuse and to regenerate a cell wall . When experiments using different growth media were compared, an inverse relationship between these abilities was observed, and a direct relationship appeared between the heterozygotes (corrected for wall regeneration) and the recombinant bacteria that were found.

J Bacteriol, 1979 Mar, 137(3), 1208 - 18
Properties of the Bacillus subtilis spore coat; Pandey NK et al.; About 70% of the protein in isolated Bacillus subtilis spore coats was solubilized by treatment with a combination of reducing and denaturing agents at alkaline pH . The residue, consisting primarily of protein, was insoluble in a variety of reagents . The soluble proteins were resolved into at least seven bands by sodium dodecyl sulfate gel electrophoresis . About one-half of the total was four proteins of 8,000 to 12,000 daltons . These were relatively tyrosine rich, and one was a glycoprotein . There was also a cluster of proteins of about 40,000 daltons and two or three in the 20,000- to 25,000-dalton range . The insoluble fraction had an amino acid composition and N-terminal pattern of amino acids very similar to those of the soluble coat proteins . A major difference was the presence of considerable dityrosine in performic acid-oxidized preparations of insoluble coats . Coat antigen including a 60,000-dalton protein not present in extracts of mature spores was detected in extracts of sporulating cells by immunoprecipitation . This large antigen turned over in a pulse-chase experiment . Antibodies to either the array of 8,000- to 12,000-dalton coat polypeptides or to the larger coat proteins reacted with this 60,000-dalton species, suggesting a common precursor for many of the mature coat polypeptides . Spore coats seem to be assembled by processing of proteins and by secondary modifications including perhaps dityrosine formation for cross-linking.

Prikl Biokhim Mikrobiol, 1979 Mar-Apr, 15(2), 314 - 7
{Pigments produced by Bacillus subtilis var . niger 16k}; Sorokulova IB et al.; Pigments produced by Bacillus subtilis var . niger 16k on tyrosine agar and histidine containing synthetic medium were isolated and identified . On the basis of a number of tests the pigments were classified as melanins.

Prikl Biokhim Mikrobiol, 1979 Mar-Apr, 15(2), 262 - 8
{Immobilization of Bacillus subtilis proteases on a styrol copolymer with maleic anhydride with covalent and coordination bonds}; Slobodianikova LS et al.; Insoluble enzymes have been obtained through an interaction of the complex preparation of proteases, protosubtilin G15x, with the sterol and maleic anhydride copolymer in aprotic dipolar solvents and as a result of formation of mixed enzyme-polymer complexes of protosubtilin G15x and the sterol and malic acid copolymer in aqueous solution in the presence of metal ions . The effect of metal ions on the yield and activity of the resultant immobilized preparation has been studied.

Mol Gen Genet, 1979 Feb 26, 170(2), 123 - 7
Host-controlled modification and restriction in Bacillus subtilis: Bsu 168-system and BsuR-system in B . subtilis 168; Ikawa S et al.; A Bsu168-specific restriction deficient (r168-) mutant of Bacillus subtilis Marburg 168 was transformed to be BsuR-specific restriction proficient (rR+) with B . subtilis R DNA as efficiently as the Bsu 168-specific restriction proficient (r168+) parental strain (hsrM+, hsdR-) . We constructed rR+ mR+ r168+ m168+ strain (ISMR 4), rR+ mR+ r168- m168+ strain (ISR 11) and rR+ mR+ r168- m168- strain (ISR 6) from strain 101 (r168+ m168+), strain 1012 (r168- m168+) and strain RM125 (r168- m168-), respectively by transformation with B . subtilis R DNA, and tested their restriction and modification activities on phage phi 105C . The results show that the sites recognized by Bsu168-specific restriction and modification enzymes and the sites recognized by BsuR-specific ones are not overlapping . We conclude that the Bsu168-modification and restriction system and the BsuR-modification and restriction system are controlled independently by two distinct sets of genes in the rR+ mR+ transformant of r168+ m168+ strain B . subtilis 168.

Mol Gen Genet, 1979 Feb 26, 170(2), 117 - 22
Mapping of genes determining nonpermissiveness and host-specific restriction to bacteriophages in Bacillus subtilis Marburg; Saito H et al.; Bacillus subtilis Marburg is nonpermissive for the multiplication of bacteriophages SP10 and phi NR2 . A permissive mutant was derived from the Marburg strain, and the genetic determinants of non-permissiveness were analyzed by PBS1 transduction . The simultaneous presence of two genes as mutant alleles, nonA and nonB, was necessary for permissiveness . The gene nonA is linked very closely to rfm (cotransfer: 95%); nonB is located between dal and purB (cotransfer of nonB and purB6 : 48%) . The genetic determinant of host-specific restriction intrinsic to the Marburg strain (hsrM) was found to be identical or very closely linked to nonB . The segregation on nonB and hsrM has never been observed in the course of transduction analysis . The mutation, hsrM1, diminishes the restriction activity, but not the host-controlled modification.

J Biol Chem, 1979 Feb 25, 254(4), 1080 - 9
Characterization of the extracellular lipase of Bacillus subtilis and its relationship to a membrane-bound lipase found in a mutant strain; Kennedy MB et al.; Bacillus subtilis CMK33 is a mutant that is more osmotically fragile than the wild type when it is converted to the protoplast form . The protoplasts of this mutant contain a membrane-bound lipase, which is not found in protoplasts of the wild type . Hydrolysis of the membrane lipid of mutant protoplasts by the lipase is the cause of their fragility . A protein found in the wild type organism specifically inhibits the lipase (Kent, C., and Lennarz, W . J . (1972) Proc . Natl . Acad . Sci . U . S . A . 69, 2793-2797) . This paper reports that cultures of both mutant and wild type cells contain an extracellular lipase which accumulates during the logarithmic phase of growth . The extracellular activity appears to be induced by a component of the growth medium . The membrane-bound lipase of the mutant has been partially purified and its properties have been compared to those of the extracellular lipase of the wild type . Their properties and sensitivity to the wild type inhibitor are similar, which suggests that the two molecules are closely related . The subcellular location of the lipase in the mutant has been investigated and compared to the location of the membrane-bound portion of the lipase inhibitor in the wild type . The lipase is located almost exclusively in the cytoplasmic membrane and not in mesosomal vesicles . In contrast, the lipase inhibitor is located in both types of membranes and is more concentrated in mesosomal vesicles . Under appropriate conditions, the appearance of new extracellular lipase activity in mutant cultures is paralleled by the loss of an equivalent amount of lipase activity from protoplasts prepared from the cells . This suggests that the membrane-bound lipase may be an intermediate in the secretion of the extracellular lipase . Because of the mutation in B . subtilis CMK33, which results in the absence of the lipase inhibitor, this intermediate can be found in protoplasts of the mutant, although it is not detectable in the wild type . Consequently, the mutant may be useful in studies of the mechanism of secretion of exoenzymes by Bacilli.

Biochim Biophys Acta, 1979 Feb 20, 551(1), 122 - 8
A monolayer study of the reaction of trinitrobenzene sulphonic acid with amino phospholipids; Bishop DG et al.; The reaction of trinitrobenzene sulphonic acid with amino phospholipids, and in particular phosphatidylethanolamine has been studied by the monolayer technique . Injection of trinitrobenzene sulphonic acid under a monolayer of amino phospholipid results in an increase in surface pressure . The rate and extent of the pressure change is greatly affected by the initial surface pressure, the fatty acid composition of the lipid, and the presence of other non-reactive lipids, especially negatively charged phospholipids . The extent of the reaction was measured with 32P-labelled phospholipids isolated from Bacillus subtilis . Only about 80% of the phosphatidylethanolamine in the monolayer could be converted to its trinitrophenyl derivative . In the presence of negatively charged phospholipids such as cardiolipin or phosphatidylglycerol, a further 20% decrease in the trinitrophenylation of phosphatidylethanolamine was found . The pressure increase occurring during trinitrophenylation could also be correlated with the extent of the reaction by comparison of the force-area curves of pure phosphatidylethanolamine, its trinitrophenyl derivative and mixtures of both compounds . The data may offer an explanation for the observation that incomplete labelling of amino phospholipids frequently occurs in natural membranes and furthermore indicate that the use of chemical labelling techniques in the study of lipid asymmetry in biological membranes must be approached with great caution.

Gene, 1979 Feb, 5(2), 87 - 91
A method for construction of specialized transducing phage rho 11 of Bacillus subtilis; Kawamura F et al.; DNA from a temperate phage rho 11 and chromosomal DNA of Bacillus subtilis 168 were digested with endonuclease EcoRI and then ligated with T4 polynucleotide ligase . The ligated DNA fragments were used to transform a lysogenic strain, B . subtilis spoA12 lys21 hisA1 leuA8 p11, and Lys+, His+ or Leu+ transformants were selected . The cells of each type were then mixed, grown and treated with mitomycin C; the induced phages were tested for abilities abilities to form plaques and to tranduce the auxotrophic marker . Various types of plaque-forming or defective phages which transduce hisA or lys marker at considerably high frequencies were thus obtained.

J Gen Microbiol, 1979 Feb, 110(2), 381 - 92
Decadent sporulation mutants of Bacillus subtilis; Balassa G et al.; In decadent sporulation mutants, sporulating populations are heterogeneous: the cells reach successive chemical and physical resistances with progressively decreasing frequencies . Each decadent mutant can be characterized by the shape and slope of the curve describing the frequency of cells resistant to various agents ('the resistance spectrum') . In some mutants the resistance spectrum decreases progressively from xylene resistance to heat resistance; in other mutants it decreases rapidly between octanol resistance and chloroform resistance . Electron microscopy showed that in two mutants the majority of the cells are blocked at stages III and IV; the number of cells that develop further to reach successive morphological stages falls off progressively . In two other mutants most cells reach stage V . Cortexless spores are also frequent . One of the decadent mutations, SpoL1, was localized between aroD and acf . The phenotype of decadent mutants is discussed in terms of sequential gene activation.

J Gen Microbiol, 1979 Feb, 110(2), 365 - 79
A Bacillus subtilis mutant requiring dipicolinic acid for the development of heat-resistant spores; Balassa G et al.; A Bacillus subtilis mutant is described which forms heat-resistant spores only in the presence of external dipicolinic acid (DPA) . The mutation, dpa-1, is localized in a new sporulation locus, linked to pyrA . The dpa-1 strain is unable to synthesize DPA but can incorporate external DPA . The amount of DPA incorporated, the frequency of heat-resistant spores and their degree of resistance are all dependent on the concentration of external DPA . Spores of dpa- 1 strains exhibit normal resistance to most chemicals, including octanol and chloroform, but not to ethanol, pyridine, phenol and trichloroacetic acid . Complete resistance to the latter group depends on DPA . DPA incorporation is slow and apparently requires an energy supply but not protein synthesis . Direct involvement of DPA in the heat-resistance of the spores is suggested . Thin sections of DPA-less spores exhibit clearly visible cytoplasmic membranes and ribosomes . These structures are absent or less visible in the core of spores obtained with added DPA.

J Gen Microbiol, 1979 Feb, 110(2), 351 - 63
Mutants of Bacillus subtilis affected in spore outgrowth; Albertini AM et al.; Six mutants of Bacillus subtilis 168 that are temperature-sensitive in spore outgrowth were isolated . The outgrowth process proceeds normally at 35 degrees C, but at the non-permissive temperature (47 degrees C) it is arrested at a specific stage characteristic for each mutant strain . The mutants are not altered in vegetative growth whether at 35 degrees C or at 47 degrees C . They were characterized for their ability to synthesize RNA, proteins and DNA during outgrowth . A mutant defective in spore germination was also isolated; less than 5% of its spores can germinate at any of the temperatures tested . The mutations were mapped by means of transduction and transformation . The isolation of a number of outgrowth mutants which map at different loci and which affect outgrowth at different times is discussed in relation to the regulation of this process.

J Antibiot (Tokyo), 1979 Feb, 32(2), 136 - 40
Preparation and antibacterial activity upon Micrococcus luteus of derivatives of iturin A, mycosubtilin and bacillomycin L, antibiotics from Bacillus subtilis; Peypoux F et al.; Methylated and acetylated derivatives of iturin A and mycosubtilin and methylated derivatives of bacillomycin L were prepared and their antibacterial activity on Micrococcus luteus was compared with the activity of the original substance . the results obtained show the importance of polar groups for the antibiotic activity of the substances of iturin group.

Biokhimiia, 1979 Feb, 44(2), 332 - 7
{Effect of pancreatic DNAse on DNA synthesis in Bacillus subtilis}; Kupriianova FG et al.; An addition of pancreatic DNAse to the cultural medium is found to stimulate DNA synthesis and proliferation of Bacillus subtilis cells . Pancreatic DNAse induces a single-stranded disruption of Bac . subtilis DNA, which may act as a mechanism of DNA synthesis increase and of the culture growth acceleration.

J Virol, 1979 Feb, 29(2), 540 - 5
Order of assembly of the lower collar and the tail proteins of Bacillus subtilis bacteriophage phi 29; Camacho A et al.; Extracts obtained after restrictive infection of Bacillus subtilis with mutants in cistron 11 of bacteriophage phi 29 are complemented in vitro by extract donors of the lower collar protein (p11) . Purified 11- heads, containing the major capsid protein (p8), the fiber protein (p8.5), the upper collar protein (p10), and the virus DNA, can be also complemented in vitro to produce infective virus . This result suggests that 11- heads are intermediates in phage phi 29 morphogenesis . The order of assembly of the lower collar protein p11 and the tail protein p9 was determined in vitro in two complementation steps . The results obtained indicate that the lower collar protein is assembled before the tail protein.

J Gen Virol, 1979 Feb, 42(2), 305 - 14
Effect of calcium ions on the infection of Bacillus subtilis by bacteriophage SF 6; Steensma HY et al.; Infection of Bacillus subtilis 168Wt by SF 6 resulted in a rapid reduction in the number of phages . This could be counteracted by the addition of calcium, barium or strontium ions . At the optimum concentration of 7.5 x 10(-2) M, the number of p.f.u . remained constant until lysis began . Although cultures of another host . B . subtilis 31 try- his-, at the end of the logarithmic growth phase produced a substance which inactivated free phages, this was not the major cause of the reduction in the numbers of p.f.u . during infection experiments at low Ca2+ concentrations . The diminution of the number of p.f.u . was therefore attributed to the fact that at least one of the steps of the lytic cycle was calcium dependent . Adsorption of SF 6 was equally effective in media containing high or low concentrations of calcium ions . Infection experiments with phages whose DNA had been labelled radioactively revealed that, at high concentrations of calcium ions, the label remained associated with the host cells until lysis commenced . At low concentrations, however, a dissociation between phage DNA and the host was found, although adsorption took place at a normal rate . From these experiments we concluded that a high concentration of calcium ions was required for the penetration of phage DNA . Similar experiments with phages whose protein coat had been labelled showed the same results, indicating that desorption of the inactivated phages occurred . Both electron microscopy and column chromatogarphy with hydroxyapatite showed that a considerable fraction of the inactivated phages had ejected their DNA into the medium . A hypothesis explaining these results is presented.

J Bacteriol, 1979 Feb, 137(2), 1028 - 30
Increased levels of dihydrofolate reductase in rifampin-resistant mutants of Bacillus subtilis; Kane JF et al.; Several independent, spontaneous rifampin-resistant mutants of Bacillus subtilis were isolated and found to have an increased resistance to trimethoprim, an inhibitor of dihydrofolate reductase . This increased resistance in the rif mutants was the result of a specific threefold increase in the activity of dihydrofolate reductase, since six other enzymes examined remained unchanged . This increased level of dihydrofolate reductase and the trimethoprim resistance were cotransformed (100%) with the rif marker . These results suggest that the RNA polymerase is altered in its recognition of the gene that specifies dihydrofolate reductase.

J Bacteriol, 1979 Feb, 137(2), 933 - 46
Possible involvement of bacterial autolytic enzymes in flagellar morphogenesis; Fein JE; Autolytic enzymes were found to be required for flagellar morphogenesis in Bacillus subtilis 168 and Bacillus licheniformis 6346 . Two previously characterized, poorly lytic, chain-forming mutants of B . subtilis 168, strains FJ3 (temperature conditional) and FJ6, each 90 to 95% deficient in the production of N-acetylmuramyl-L-alanine amidase and endo-beta-N-acetylglucosaminidase, were observed to be nonmotile at 35 degrees C in a variety of liquid and semisolid meida . In contrast, cells of the isogenic wild-type strain were motile and fully separated . Electron microscopy revealed the complete absence of flagella on the mutant cells . Similar observations were made with another poorly lytic strain of B . subtilis 168 (Nil5) and with two poorly lytic, phosphoglucomutase-deficient mutants of B . licheniformis 6346 (MH-3, MH-5) . In minimal media lacking galactose (restrictive conditions), the B . licheniformis mutants failed to form flagella, or had serious abnormalities in flagellar morphogenesis and motility . Under permissive conditions, mutants FJ3 (grown at 17 degrees C) and MH-5 (grown with addend galactose) showed increased autolytic activities, grew in the dechained form, and regained their capacities to synthesize functional flagella . Examination of several classes of spontaneous revertants derived from the various mutant strains further demonstrated a close relationship between autolysin acttivity and flagellation in the two Bacillus spp.

J Bacteriol, 1979 Feb, 137(2), 773 - 8
Purification and properties of the manganese-dependent phosphoglycerate mutase of Bacillus subtilis; Watabe K et al.; Phosphoglycerate mutase of Bacillus subtilis was purified to apparent homogeneity . It specifically required manganese ions for stability and activity, but it does not need 2,3-diphosphoglycerate as cofactor; the Km for Mn2+ is about 4.5 micrometer . Enzyme activity was inhibited by heavy-metal ions, 2,3-butanedione, and sulfhydryl agents . The mutase has a molecular weight of about 74,000 as shown by Sephadex gel filtration and by acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate; it consisted of one polypeptide.

Mol Gen Genet, 1979 Jan 10, 168(2), 165 - 72
The genome of Bacillus subtilis phage SPP1: the arrangement of restriction endonuclease generated fragments; Ratcliff SW et al.; SPP1 DNA was cleaved by the restriction endonucleases, BglI, BglII, EcoRI, KpnI, SmaI, and SalI . The molecular weights of the DNA fragments obtained by single enzyme digestion or by consecutive digestion with two enzymes were determined by electron microscopic measurements of contour length and by gel electrophoresis . The major fragments from the six digests could be ordered to give a consistent restriction map of SPP1 . The electropherograms of several digests indicated that certain fragments occurred in less than stoichiometric amounts or were heterogeneous in size . Such bands carried a major part of radioactivity, when SPP1 DNA was terminally labelled with P32 prior to degradation by restriction enzymes . These results, and studies of the effect of exonuclease III treatment on restriction enzyme patterns define the terminal restriction fragments . All data obtained support the conclusion drawn in the preceding paper (Morelli et al., 1978 b) that the SPP1 genome is terminally redundant and partially circularly permuted.

Biochemistry, 1979 Jan 9, 18(1), 198 - 202
Extracellular labeling of growing secreted polypeptide chains in Bacillus subtilis with diazoiodosulfanilic acid; Smith WP et al.; Studies of the mechanism of protein secretion in a Gram-positive bacterium, Bacillus subtilis, yielded results very similar to those previously obtained with a Gram-negative organism: nascent chains protruding from protoplasts could be labeled extracellularly; the labeled chains could be recovered on polysomes isolated from the membrane--polysome fraction; they could be released by puromycin, low Mg2+, or chain completion; the completed chains include a known secreted protein (alpha-amylase); and their ribosomes appear to be attached to membrane solely by their nascent chains . The reagent used for extracellular labeling, {1252}diazoiodosulfanilic acid, yielded severalfold more specific labeling of the nascent chains (7--10% of the total cellular labeling and one-fourth to one-third of that of the membrane--polysome fraction) than was obtained earlier with another nonpenetrating reagent.

Mol Gen Genet, 1979 Jan 5, 168(1), 111 - 5
High frequency transformation of Bacillus subtilis protoplasts by plasmid DNA; Chang S et al.; A highly efficient method for transformation of Bacillus subtilis by plasmid DNA is reported . The procedure, which involves polyethylene glycol-induced DNA uptake by protoplasts and subsequent regeneration of the bacterial cell wall, yields up to 80% transformants with an efficiency of 4 x 10(7) transformants per microgram of supercoiled DNA . Plasmids constructed by in vitro ligation or endonuclease-generated fragments of linear plasmid DNA can also transform PEG-treated protoplasts, but at a lower frequency.

Mol Gen Genet, 1979 Jan 2, 167(3), 251 - 8
Characterization of plasmid transformation in Bacillus subtilis: kinetic properties and the effect of DNA conformation; Contente S et al.; Transformation of competent cells of Bacillus subtilis with antibiotic resistance plasmid DNA has shown that (a) competence for plasmid and chromosomal DNA develops with similar kinetics; (b) DNA linearized with a variety of restriction endonucleases does not transform; (c) CCC plasmid DNA is inactivated for transformation by a single nick; (d) T4 ligase restores transforming activity to both nicked and linearized DNA; (E) CCC relaxed DNA is fully active in transformation; (f) the DNA concentration-dependence of plasmid transformation is first order; and (g) plasmid transformation proceeds with a low efficiency, requiring the uptake of 10(3) to 10(4) DNA molecules per transformant . Based on this information, a model for the processing of chromosomal, plasmid and transfecting DNA is proposed.

Pharmazie, 1979, 34(9), 548 - 51
{Interactions between macromolecular adjuvants and drugs . Part 18: The binding behaviour of sodium carboxymethylcellulose and other macromolecules towards streptomycin sulphate (author's transl)}; Keipert S et al.; Unlike the other macromolecules under investigation (polyvinylpyrrolidone, methylcellulose, polyvinyl alcohol), sodium carboxymethylcellulose acts on streptomycin sulphate, which leads to isolable precipitates . By equilibrium dialysis in electrolyte-free and electrolyte-containing solutions, the interaction was characterized in more detail and described by Donnan curves . The associates (which contained, on an average, 40% of streptomycin) proved to be resistant to light and air, and stable to thin-layer chromatography . Microbiological assays on the test strain Bacillus subtilis SG119 revealed a decrease in activity of the streptomycin-sodium carboxymethylcellulose associates compared to pure streptomycin (tested in the solid form, using rice granulate as a carrier substance) . In contrast, the associate was as efficient as the antibiotic when dissolved in a concentrated solution of sodium chloride.

Folia Microbiol (Praha), 1979, 24(6), 462 - 72
A psychrophilic strain Actinoplanes sp . 220; Ricicova A et al.; The strain designated Actinoplanes sp . 220 differed in its characteristics from other strains of the genus Actinoplanes listed in Bergey's Manual (1974) . The strain belongs to psychrophilic culture growing within the range of 0-30 degrees C . The optimal temperature for growth on yeast--malt agar is 10-23 degrees C . Cultures transferred at 23 and 28 degrees C differed in morphological and physiological properties, enzyme activity and pigmentation in standard media . Submerged culture transferred at 28 degrees C inhibited growth of Bacillus subtilis ATCC 6633 and ATCC 9945 . LL-2,6-Diaminopimelic acid was chromatographically detected in the submerged mycelium of this culture . This compound was not found in the mycelium of the original culture transferred at 23 degrees C . The cultures did not substantially differ in the composition of other amino acids contained in larger quantities in the mycelium.

Microbios, 1979, 24(96), 113 - 22
Measurement of pyrimidine dimers in spheroplasts of Bacillus subtilis; Hadden CT; A method is described for making spheroplasts of Bacillus subtilis which are permeable to exogenous enzymes . Conditions are described for measuring small numbers of pyrimidine dimers in the DNA of UV-irradiated cells by use of a partially purified Micrococcus luteus extract containing an enzyme specific for pyrimidine dimers . The system will detect as few as 10-12 pyrimidine dimers per genome.

J Bacteriol, 1979 Jan, 137(1), 82 - 91
Characterization of pyrimidine-repressible and arginine-repressible carbamyl phosphate synthetases from Bacillus subtilis; Paulus TJ et al.; The number and properties of carbamyl phosphate synthetases in Bacillus subtilis have been uncertain because of conflicting genetic results and instability of the enzyme in extracts . The discovery of a previously unrecognized requirement of B . subtilis carbamyl phosphate synthetases for a high concentration of potassium ions for activity and stability permitted unequivocal demonstration that this bacterium elaborates two carbamyl phosphate synthetases . Carbamyl phosphate synthetase A was shown to be repressed by arginine, to have a molecular weight of about 200,000, and to be coded for by a gene that maps near argC4 . This isozyme was insensitive to metabolites of the arginine and pyrimidine biosynthetic pathways . Carbamyl phosphate synthetase P was found to be repressed by uracil, to have a molecular weight of 90,000 to 100,000, and to be coded for by a gene that maps near the other pyr genes . This isozyme was activated by phosphoridine nucleotides . Other kinetic properties of the two isozymes were compared . Bacillus thus resembles eucaryotic microbes in producing two carbamyl phosphate synthetases, rather than the enteric bacteria, which produce a single carbamyl phosphate synthetase.

Zentralbl Bakteriol Naturwiss, 1979, 134(4), 352 - 9
Subtilopeptidase A produced by Bacillus subtilis PR-70 . II . Kinetic behaviour of immobilized enzyme; Attia RM et al.; The kinetic behaviour of immobilized subtilopeptidase A was investigated . The enzyme was obtained from a local isolate of B . subtilis PR-70 . Using different inorganic supports, Amberlite CG-50 was superior in this respect . It gave 97.8% adsorption, followed by silica gel GC . The values of K and K2 for the rate of enzyme catalyzed being 8.75 and 2.06, respectively . The behaviour of v against Et is the same as v against St . Michaelis' constant was determined using different methods . The average of Km value and Vmax were 0.0094 and 0.95, respectively . Studying how v behaves when St is varied while Et is constant, two active site per enzyme molecule and auto-inhibition of enzyme by its own substrate were observed . Comparing kinetic parameters of a soluble and insoluble subtilopeptidase A showed that Km decreased from 0.016 to 0.0094, while Vmax increased from 0.71 to 0.95, respectively . This indicated that when subtilopeptidase was bound to Amb . GC-50, a case of partially non-competitive inhibition occurred . The recovery of enzymatic activity in the water insoluble subtilopeptidase A is 12.8 per cent.

Microbiol Immunol, 1979, 23(8), 727 - 34
Growth and sporulation of auxotrophs of Bacillus subtilis in a medium with limited nutrients; Nishihara T; Growth and sporulation were examined for 30 auxotrophs of Bacillus subtilis in a chemically defined medium with suboptimal amounts of nutrients . All strains except for some adenine-requiring mutants could not overtake sporulation stage II when amino acids, vitamins, or bases were limited, whereas they sporulated fairly well without limitation . Abnormal structures, a cell with thickened cell wall and a cell with several refractile bodies, were found in some strains after the vegetative growth stopped.

Mol Gen Genet, 1979, 177(1), 23 - 9
Chromosomal mutations causing resistance to tetracycline in Bacillus subtilis; Williams G et al.; We have isolated, after ethylmethanesulfonate mutagenesis, several chromosomal mutations causing resistance to tetracycline in Bacillus subtilis . These mutations fall into two classics, tetA and tetB . 30 S ribosomal protein S10 shows an altered mobility on two-dimensional acrylamide gels in cells bearing the former type of mutation . Ribosomes from these cells show elevated levels of resistance to tetracycline in vitro as measured by polyuridine dependent polyphenylalanine synthesis . The tetA locus maps adjacent to the tuf gene in the B . subtilis ribosomal protein gene cluster . Cells with the tetB mutation do not show any altered ribosomal protein, and their ribosomes are as sensitive, in vitro, to tetracycline as ribosomes isolated from wild type cells . The tetB mutation has been mapped proximal to cysA14.

Ultramicroscopy, 1979, 4(4), 395 - 412
Low-dose electron image reconstruction of negatively stained contractile phage sheath from Bacillus subtilis (PBS-Z); Cremers AF et al.; The structure of the contractile sheath of the defective phage from B . subtilis (PBS-Z) has been investigated by low-dose electron microscopy and image reconstruction . The extended and contracted sheath particles were imaged by means of two negative stains which consisted of uranyl- and phosphotungstate-containing solutions of a pH of 4.2 and 7.0 respectively . Images of identical parts of the same type of specimen were recorded at a total electron dose of 80 C/m2 (5 electrons/A2) and 4 x 10(3) C/m2 (250 electrons/A2) . The low-dose reconstructions of the extended and contracted sheath structure in the two stains show good correspondence and made it possible to draw the following structural conclusions . The sheath protein in both types of structure has an elongated shape, and in both structures the long molecular axis lies in a plane perpendicular to the helical sheath axis . The orientation of the protein in the extended and contracted sheath is different; the long axes differ by about 35 degrees in orientation . The reconstructions did not permit conclusions about different conformational states of the protein in both structures . These data, together with the packing parameters of the protein subunits in the contractile sheath {1}, form the complete structural analysis of this biological structure by electron microscopy . The radiation damage effects which have been monitored in analyzing image pairs to the full extent may be summarized as follows . (1) Diameters of the sheath structure increase, which indicate flattening . (2) There is no loss in resolution, and layerline altitudes of the Fourier-transformed images do not change . (3) Uranyl stain behaves differently compared to phosphotungstate . In both negative stains the structural noise level increases upon irradiation as follows from the increase in phase residuals of the digital layerline data . In uranyl-stained images also more aperiodic noise appears . (4) The Fourier amplitudes of the principal layerline maxima shift towards lower spatial frequencies; phases of corresponding maxima generally remain constant . This pattern is more pronounced in the extended sheath data; there is no rationale describing these positional shifts . Moreover, in the case of contracted sheath the amplitudes of Fourier components also change more in absolute value . Therefore the damage effects also seem to depend on the type of structure embedded in the stain . (5) In the reconstructed images these radiation effects create artificial stain-excluded volumes of a type and at a radius which depend on the stain and structure.

Microbios, 1979, 24(95), 29 - 39
Comparison of three procedures for isolating DNA from bacteria; Amundesn SK et al.; Three methods employing chloroform-isoamyl alcohol (CI), phenol, or enzymes, were evaluated for isolating DNA from Escherichia coli, Bacillus subtilis, and Arthrobacter globiformis . For the amounts of reagents employed at optimum conditions in the CI and phenol procedures, 0.4-0.9 mg of DNA/g wet weight of cells was isolated . Using the enzymatic procedure, approximately twice as much DNA was isolated . DNA isolated by the CI procedure contained 0.03-0.09% protein and 0.08-0.12% RNA . DNA isolated by the phenol procedure contained 0.02-0.05% protein and 2.2-2.6% RNA . DNA isolated by an enzymatic procedure, which is described in detail, contained 32.2-45.7% protein and 0.3-0.6% RNA . DNA isolated by all three procedures are double-stranded and at least 10(6) in molecular weight, as suggested by data from thermal transition analyses and transformations . These data emphasize that the desired characteristics of DNA for experimental purposes must be considered in selecting an isolation procedure.

Folia Microbiol (Praha), 1979, 24(5), 373 - 5
Exoprotease production by sporogenous and asporogenous mycobacillin non-producer mutants of Bacillus subtilis; Bose R et al.; Mycobacillin non-producers, whether sporogenous or asporogenous, possess less exoprotease, but effective exoprotease producers are not always good mycobacillin yielders . There might exist a minimum level of exoprotease formation for elaboration of mycobacillin.

Genetika, 1979, 15(3), 594 - 604
{Production and study of Bacillus subtilis mutants for genes involved in nucleoside catabolism}; Rumiantseva EV et al.; By means of selection for a low thymine requirement the mutants fo thymine auxotrophs for deoxyriboaldolase (dra) and phosphodeoxyribomutase (drm) genes were obtained . Besides the mutants for pyrimidinenucleoside phosphorylase gene (pdp) were olso isolated using selection on the fluorodeoxyuridine resistance . The latter enzyme provides for pyrimidine nucleosides catabolism (thymidine, uridine) in Bacilli, as well as the conversion of exogenous thymine to thymidine in thymine auxotrophs . The data obtained when studying the deo-enzymes activities in various types of the mutants and also under the condition of induction by thymidine and acetoaldehyde are in accordance with the assumption that deoxyriboso-5-phosphate is an inductor of the deo-enzymes in Bacillus subtilis . The genes dra and pdp were tightly linked as it had been shown by the transformation experiments; in contrast, no linkage was revealed between dra and drm or pdp and drm . A secondary mutation (adn), not linked with dra and blocking the ability of bacteria to catabolise adenosine (purine nucleoside phosphorylase activity remains constant) was found in some dra-mutants.

Zentralbl Bakteriol Naturwiss, 1979, 134(3), 275 - 81
Subtilopeptidase A produced by Bacillus subtilis PR-70 . I . Kinetic behaviour of solubilized enzymes; Attia RM et al.; The kinetic behaviour of subtilopeptidase A was investigated . The enzyme was obtained from a local isolate of B . subtilis PR-70 . The rate of enzyme catalyzed conversion of substrate to product is directly proportional to the enzyme concentration, v = K(E) . Michaelis constant was determined using different methods . The average of Km value is equal to 0.01615 . The Vmax and Et were determined being 0.71 and 1.467, respectively, using KUNITZ's casein digestion method . The enzymeconcentration involved in the reaction system is equal to 97.8% . A trial to calculate the molecular weight and the number of active groups were discussed.

Biochimie, 1979, 61(3), 385 - 91
Characterization of an inhibitor of the intracellular protease from Bacillus subtilis; Millet J et al.; A specific inhibitor of intracellular serylprotease from Bacillus subtilis has been isolated from both growing and sporulating cells . Like other protease inhibitors isolated from eukaryotic cells, the inhibitor from B . subtilis is a thermostable protein . A purification method is described . The molecular weight estimated by Biogel filtration and SDS gel electrophoresis is about 15,500 . Both proteolytic and esterolytic activities of intracellular protease are equally sensitive to inhibition . With azocoll or Z-tyrosine p-nitrophenylester as substrates, noncompetitive inhibition patterns are observed . The inhibitor has no effect on the proteolytic or esterolytic activities of the extracellular serylprotease . A similar thermostable inhibitor is also present in Bacillus megaterium.

Biochimie, 1979, 61(1), 93 - 100
On the nature of tetracycline resistance in Bacillus subtilis mediated by the plasmid pT 127; Fargette F et al.; The nature of tetracycline resistance was studied in a strain of Bacillus subtilis carrying the plasmid pT 127 in comparison with the parental strain . The resistance has been shown to be inducible in both strains upon exposure to subinhibitory concentrations of tetracycline . No modification of the protein-synthesizing activity of the ribosomes or intracellular inactivation of the antibiotic was observed in both strains . Accumulation of labeled tetracycline in B . subtilis was found to be particularly low in the wild-type strain, compared to other bacterial species, with concentration gradients of only 2 to 3 fold . From the kinetics obtained it is likely that the permeation of the antibiotic does not correspond to an active process in B . subtilis . A fairly good correlation was established between the level of resistance obtained after induction or by the presence of the plasmid pT 127 and a decrease in the binding capacity of the cell for the antibiotic.

J Virol, 1979 Jan, 29(1), 61 - 8
Deoxythymidine nucleotide metabolism in Bacillus subtilis W23 infected with bacteriophage SP1Oc: preliminary evidence that dTMP in SP10c DNA is synthesized by a novel, bacteriophage-specific mechanism; Markewych O et al.; Despite the fact that mature SP10c DNA contains dTMP, the acid-soluble fraction of infected cells contained no dTTP during the interval of phage replication . However, infected cells contained normal cellular levels of dATP, dGTP, and dCTP . Upon infection of deoxythymidine-starved Bacillus subtilis M160 (a deoxythymidine-requiring mutant of B . subtilis W23), mature phage DNA with a normal dTMP content was made . SP10c codes for an enzyme that seems to catalyze the tetrahydrofolate-dependent transfer of 1-carbon fragments to the 5 position of dUMP . The transfer of 1-carbon fragments is not accompanied by oxidation of tetrahydrofolage to dihydrofolate, implying that the enzyme in question is not a dTMP synthetase . It is proposed that dTMP in mature SP10c DNA is derived by the postreplicational modification of some other nucleotide and not by the direct incorporation of dTTP into DNA.

Gene, 1979 Jan, 5(1), 1 - 7
Unusual base sequence arrangement in phage phi 29 DNA; Ito J et al.; Susceptibility of Bacillus subtilis phage phi 29 DNA to 34 different restriction endoculceases was determined . Three enzymes, BglI, XbaI and BstEII, were found to cleave phi 29 DNA only once at specific sites . The sites of these single cleavages have been mapped . Thirteen enzymes did not cut phi 29 DNA . phi 29 HindIII DNA fragments inserted into pBR313 plasmid and propagated in Escherichia coli, were resistant to these restriction endonucleases . This result suggests that the insusceptibility is due to the absence of the nucleotide sequences on phi 29 recognized by the enzymes, and not to the presence of modified nucleotides.

Mikrobiologiia, 1979 Jan-Feb, 48(1), 93 - 8
{Experimental verification of the results of the mathematical modeling of the process of formation of alpha-amylase-nonproductive mutants of Bacillus subtilis under continous cultivation}; Fencl Z et al.; When Bacillus subtilis produces alpha-amylase in the course of continuous cultivation, it is difficult to maintain the activity at a constant level . This may be due to the formation of nonproductive mutants . Individual cells in the population have been analysed in the course of the continuous process . The composition of the population changes depending on time and the composition of the growth medium . Semisynthetic media cause selection of mutants which synthesize the enzyme at a low rate . In contrast, complex media which are more enriched in the sources of carbon and nitrogen induce accumulation of mutants with a high activity.

J Bacteriol, 1979 Jan, 137(1), 635 - 43
Plasmid copy number control: isolation and characterization of high-copy-number mutants of plasmid pE194; Weisblum B et al.; A plasmid, pE194, obtained from Staphylococcus aureus confers resistance to macrolide, lincosamide, and streptogramin type B ("MLS") antibiotics . For full expression, the resistance phenotype requires a period of induction by subinhibitory concentrations of erythromycin . A copy number in the range of 10 to 25 copies per cell is maintained during cultivation at 32 degrees C . It is possible to transfer pE194 to Bacillus subtilis by transformation . In B . subtilis, the plasmid is maintained at a copy number of approximately 10 per cell at 37 degrees C, and resistance is inducible . Tylosin, a macrolide antibiotic which resembles erythromycin structurally and to which erythromycin induces resistance, lacks inducing activity . Two types of plasmid mutants were obtained and characterized after selection on medium containing 10 microgram of tylosin per ml . One mutant class appeared to express resistance constitutively and maintained a copy number indistinguishable from that of the parent plasmid . The other mutant type had a 5- to 10-fold-elevated plasmid copy number (i.e., 50 to 100 copies per cell) and expressed resistance inducibly . Both classes of tylosin-resistant mutants were shown to be due to alterations in the plasmid and not to modifications of the host genome.

J Bacteriol, 1979 Jan, 137(1), 391 - 6
DNA repair in Bacillus subtilis: excision repair capacity of competent cells; Yasbin RE et al.; Competent Bacillus subtilis were investigated for their ability to support the repair of UV-irradiated bacteriophage and bacteriophage DNA . UV-irradiated bacteriophage DNA cannot be repaired to the same level as UV-irradiated bacteriophage, suggesting a deficiency in the ability of competent cells to repair UV damage . However, competent cells were as repair proficient as noncompetent cells in their ability to repair irradiated bacteriophage in marker rescue experiments . The increased sensitivity of irradiated DNA is shown to be due to the inability of excision repair to function on transfecting DNA in competent bacteria . Furthermore, competent cells show no evidence of possessing an inducible BsuR restriction system to complement their inducible BsuR modification enzyme.

J Bacteriol, 1979 Jan, 137(1), 35 - 43
Cell wall teichoic acid as a reserve phosphate source in Bacillus subtilis; Grant WD; Although exponential growth of Bacillus subtilis 168 in a phosphate-limited medium halted with the exhaustion of inorganic phosphate, the bacteria continued to grow at a slower rate for a further 3 to 4 h at 37 degrees C . This postexponential growth in the absence of an exogenous phosphate supply was accompanied by a loss of teichoic acid from the cell walls of the bacteria . Quantitative analysis of walls and culture fluids showed that the phosphate loss from the walls could not be accounted for by an increase in phosphate-containing compounds in the medium, which implied that the cells were using their own wall teichoic acids to supply phosphate necessary for growth . Addition of exogenous teichoic acid to phosphate-starved cultures resulted in stimulation of growth and in the simultaneous disappearance of teichoic acid phosphate from the medium . It is proposed that teichoic acids, which can contain more than 30% of the total phosphorus of exponential-phase cells, can be used as a reserve phosphate source when the bacteria are starved for inorganic phosphate.

J Bacteriol, 1979 Jan, 137(1), 327 - 31
Control of teichoic acid synthesis during phosphate limitation; Glaser L et al.; The synthesis of teichoic acids was examined in Bacillus subtilis Marburg grown under conditions of phosphate limitation . The results indicate that the inhibition of polyglycerolphosphate synthesis observed under these conditions is the result of two processes . The first process is reversible and is independent of new protein synthesis; the second process is irreversible and requires the synthesis of new protein . During growth, under conditions of phosphate limitation, there is a slow decrease in the level of CDP glycerol pyrophosphorylase activity which is by itself not sufficient to account for the decrease in the rate of polyglycerolphosphate synthesis.

J Bacteriol, 1979 Jan, 137(1), 213 - 20
Inhibition of Bacillus subtilis growth and sporulation by threonine; Lamb DH et al.; A 1-mg/ml amount of threonine (8.4 mM) inhibited growth and sporulation of Bacillus subtilis 168 . Inhibition of sporulation was efficiently reversed by valine and less efficiently by pyruvate, arginine, glutamine, and isoleucine . Inhibition of vegetative growth was reversed by asparate and glutamate as well as by valine, arginine, or glutamine . Cells in minimal growth medium were inhibited only transiently by very high concentrations of threonine, whereas inhibition of sporulation was permanent . Addition of threonine prevented the normal increase in alkaline phosphatase and reduced the production of extracellular protease by about 50%, suggesting that threonine blocked the sporulation process relatively early . 2-Ketobutyrate was able to mimic the effect of threonine on sporulation . Sporulation in a strain selected for resistance to azaleucine was partially resistant . Seventy-five percent of the mutants selected for the ability to grow vegetatively in the presence of high threonine concentrations were found to be simultaneously isoleucine auxotrophs . In at least one of these mutants, the threonine resistance phenotpye could not be dissociated from the isoleucine requirement by transformation . This mutation was closely linked to a known ilvA mutation (recombination index, 0.16) . This strain also had reduced intracellular threonine deaminase activity . These results suggest that threonine inhibits B . subtilis by causing valine starvation.

J Bacteriol, 1979 Jan, 137(1), 189 - 96
Carbon and nitrogen repression of arginine catabolic enzymes in Bacillus subtilis; Baumberg S et al.; Specific activities of arginase and ornithine aminotransferase, inducible enzymes of arginine catabolism in Bacillus subtilis 168, were examined in cells grown with various carbon and nitrogen sources . Levels of these enzymes were similar in arginine-induced cultures whether glucose or citrate was the carbon source (in contrast to histidase), suggesting that carbon source catabolite repression has only limited effect . In media with combinations of nitrogen sources, glutamine strongly repressed induction of these enzymes by proline or arginine . Ammonium, however, only repressed induction by proline and had no effect on induction by arginine . These effects correlate with generation times in media containing these substances as sole nitrogen sources: growth rates decreased in the order glutamine-arginine-ammonium-proline . Similar phenomena were observed when glutamine or ammonium were added to arginine- or proline-grown cultures, or when arginine or proline were added to glutamine- or ammonium-grown cultures . In the latter cases, an additional feature was apparent, namely a surprisingly long transition between steady-state enzyme levels . The results are compared with those for other bacteria and for eucaryotic microorganisms.

J Bacteriol, 1979 Jan, 137(1), 124 - 8
Effect of 6-(p-hydroxyphenylazo)-uracil on the homologous and heterologous transduction processes in Bacillus subtilis; Canosi U et al.; We have studied the effect of 6-(p-hydroxyphenylazo)-uracil on the recombination processes that operate in the homologous and heterologous transduction mediated by PBS1 and SP10 phages of Bacillus subtilis . The results obtained demonstrate that the process of heterologous genetic exchange is sensitive to this compound, whereas the homologous process is not . The present data, along with those of our previous work (U . Canosi, A . G . Siccardi, A . Falaschi, and G . Mazza, J . Bacteriol . 126:108--121, 1976), suggest that the DNA polymerase III is involved in the recombination process that operates in transformation and heterologous transduction, whereas homologous transduction follows a partially independent pathway not involving this protein.

Prikl Biokhim Mikrobiol, 1979 Jan-Feb, 15(1), 57 - 62
{Effect of sources of carbon, nitrogen, and phosphorus on the synthesis of proteases from Bacillus subtilis cultures}; Kaluniants KA et al.; The paper gives data on the influence of different sources of carbon, nitrogen and phosphorus on the accumulation of biomass and synthesis of neutral protease by Bacillus subtilis str . 103 . The highest proteolytic activity was obtained on the medium containing maltose or hydrolyzed starch as a carbon source, monopotassium phosphate as a phosphorus source, and ammonium sulphate as a nitrogen source . The enzyme activity increased upon an addition of albumin, peptone or casein . A study of the amino acid effect showed that methionine, isoleucine and valine stimulated the protease synthesis to the greatest extent.

Biochim Biophys Acta, 1978 Dec 21, 521(2), 719 - 25
Asymmetric transcription during post-germinative development of Bacillus subtilis spores . II . Hybrid competition analyses; Setoguchi Y et al.; Hybrid-competition analyses were done to estimate the relatedness of 3H-labeled mRNA species synthesized during spore germination and log-phase growth . The competitions showed that early in the germination process 10--15 and 1--3% of the RNA transcribed from the H and from the L strand, respectively, were unique and absent during log-phase growth . At later stages, the amounts of the germination-specific H transcripts decreased more rapidly than the L transcripts . The competitions with pulse-labeled log-phase RNAs showed that vegetative genes were transcribed more rapidly from the H strand than from the L strand . Most of the results could be correlated with the observed decrease in the H/L asymmetry ration during spore germination.

Biochim Biophys Acta, 1978 Dec 21, 521(2), 708 - 18
Asymmetric transcription during post-germinative development of Bacillus subtilis spores; Margulies L et al.; The relative transcription from L and H strands of Bacillus subtilis DNA during consecutive stages of spore outgrowth was determined and compared to the transcription pattern during log-phase growth of vegetative cells . Pulses of {3H} uridine were administered during early, middle and late outgrowth phases of germination and the RNAs isolated . The asymmetry ratio of H/L as determined by hybridization at saturating RNA/DNA inputs showed a gradual decrease . During the period studied (10-90 and 90-160 min post-induction), about 50 and 35%, respectively, of the radioactive RNA consisted of ribosomal RNA transcripts . The decrease in the H/L asymmetry ratio was due predominantly to the appearance and accumulation of L strand transcripts and not to either changes in the quantity of H strand transcripts nor to fluctuation in the rate of rRNA synthesis.

Biochim Biophys Acta, 1978 Dec 21, 521(2), 484 - 92
Transformation in Bacillus subtilis with nitrogen mustard crosslinked DNA . Effect on cotransformation and mutation frequencies; Scheinbach S et al.; Bacillus subtilis DNA was treated in vitro with nitrogen mustard and the crosslinked molecules were purified, after alkali denaturation, by hydroxyapatite chromatography . When tested for the ability to transform the trpC2-hisB2 segment, these molecules exhibited a decrease in the cotransformation index (r) as compared to native or renatured DNA . The decrease in r was not accompanied by an increase in mutagenicity.

Arch Microbiol, 1978 Dec 20, 119(3), 287 - 93
On the appearance of Bacillus subtilis intracellular serine protease in the cell membrane and culture medium . Comparison of the enzyme and other Bacillus subtilis serine proteases; Strongin AY et al.; While about 80% of the cell-bound intracellular serine protease of Bacillus subtilis A-50 have been recovered in the soluble fraction upon disruption of cells, the rest of the enzyme was found to be associated with the membrane fraction . Soluble cytoplasmic intracellular serine protease, as well as membrane-bound serine protease liberated by non-ionic detergent treatment, have been isolated in a pure state and shown to be identical . The same protease might also be found extracellularly, due presumably to cell lysis or altered membrane permeability . Intracellular serine protease of Bacillus subtilis A-50 was clearly related to Bacillus subtilis serine proteases W1 and bacillopeptidase F described as extracellular enzymes.

Biochem J, 1978 Dec 15, 176(3), 639 - 47
Inhibition of bacterial transport by uncouplers of oxidative phosphorylation . Effects of pentachlorophenol and analogues in Bacillus subtilis; Nicholas RA et al.; Analogues of the potent uncoupler of oxidative phosphorylation pentachlorophenol were tested as inhibitors of proline and glycine transport by Bacillus subtilis . These analogues included less highly substituted chlorophenols and pentachlorothiophenol . Like pentachlorophenol, they are non-competitive inhibitors of proline transport and uncompetitive inhibitors of glycine transport . However, the less highly substituted chlorophenols are weaker acids than pentachlorophenol and also weaker inhibitors . Analysis indicated that the anionic form of the uncouplers is the inhibiting species . Pentachlorothiophenol, a water-insoluble anion, is also a potent inhibitor . These results support previous studies that concluded that uncouplers of oxidative phosphorylation inhibit amino acid transport by binding at specific sites on proteins, the free energy of interaction stabilizing 'unproductive' conformations . Such specific interactions of uncoupler with protein are probably commonplace.

J Biol Chem, 1978 Dec 10, 253(23), 8559 - 63
Purification and properties of a Bacillus subtilis endonuclease specific for apurinic sites in DNA; Inoue T et al.; An endonuclease which hydrolyzes depurinated DNA has been purified from extracts of Bacillus subtilis cells . The endonuclease is a monomeric protein and has a molecular weight of around 56,000 . The enzyme is specific for apurinic sites in double-stranded DNA, has a pH optimum at 8.0, and is slightly stimulated with 50 mM NaCl but completely inhibited with 500 mM NaCl . It requires no divalent cations and is insensitive to EDTA; it has no associated exonuclease . These properties are very similar to those of Escherichia coli endonuclease IV, which is also insensitive to EDTA and has no exonuclease activity, and very different from those of the main endonuclease for apurinic sites (endonuclease IV) of the same bacterium.

J Biol Chem, 1978 Dec 10, 253(23), 8518 - 25
Bacillus subtilis bacteriophage PBS2-induced DNA polymerase . Its purification and assay characteristics; Hitzeman RA et al.; The DNA polymerase induced by Bacillus subtilis bacteriophage PBS2 (whose DNA contains uracil instead of thymine) has been purified and characterized for its specificity . The enzyme requires a high ionic strength for optimal stability and activity and is sensitive to various anions and to sulfhdryl reagents . Both dUTP and dTTP are incorporated efficiently as substrates and are competitive inhibitors at the same active site . The apparent Km and Ki values are about 6 micrometers for dTTP and 15 micrometers for dUTP, when denatured, uracil-containing B . subtilis or salmon sperm DNA (3.9 micrometers for dUTP and 2.6 micrometers for dTTP) . The PBS2 enzyme works best on denatured DNA, on double-stranded DNA activated by DNase to produce gaps, or on primed homopolymeric DNA . Using denatured DNA preparations of average molecular weight 6.2 million, the apparent Km values are 270 micrograms/ml for B . subtilis DNA and 360 micrograms/ml for PBS2 DNA; the Vmax value for denatured PBS2 DNA containing uracil is 7-fold greater than that for denatured B . subtilis DNA containing thymine . However, lower molecular weight DNAs have 10-fold lower apparent Km values and show similar Vmax values for both B . subtilis and PBS2 DNAs . Thus, the PBS2 phage-induced DNA polymerase (which likely replicates only uracil-containing phage DNA using dUTP in vivo) has little selectivity for uracil- versus thymine-containing deoxyribonucleotides or DNA in vitro.

J Biol Chem, 1978 Dec 10, 253(23), 8526 - 32
Characterization of the Bacillus subtilis bacteriophage PBS2-induced DNA polymerase and its associated exonuclease activity; Hitzeman RA et al.; The DNA polymerase induced by Bacillus subtilis bacteriophage PBS2 has a Stokes radius of 7.2 in buffers of high ioninc strength, suggesting a molecular weight in the range 145,000 to 195,000 . The polypeptide bands observed on gel electrophoresis in dodecyl sulfate have apparent molecular weights of 78,000 and 69,000 (and possibly another 27,000) in equimolar amounts . In buffers of low ionic strength, the enzyme appears to form large aggregates and even precipitates, with about 90% loss of activity . A nuclease activity co-purifies with the PBS2 DNA polymerase and shows similar responses to changes in pH, MgCl2, N-ethylmaleimide, temperature, and dextran sulfate levels . The nuclease produces deoxyribonucleoside 5'monophosphates from denatured DNA containing thymine or uracil . No endonuclease activity is detectable on supercoiled DNA . The inhibition of nuclease activity by added deoxyribonucleoside triphosphates, the DNA-dependent turnover of triphosphates, to free monophosphates during DNA polymerization, the inhibition of nuclease activity by 3'-phosphates on the DNA template-primer, and the pattern of digestion of 5'-{32P}phosphate-labeled DNA all indicate that the PBS2 DNA polymerase-associated hydrolytic activity is a 3' leads to 5'-exonuclease.

J Gen Microbiol, 1978 Dec, 109(2), 225 - 36
Linkage analysis in Dictyostelium discoideum using multiply marked tester strains: establishment of linkage group VII and the reassessment of earlier linkage data; Ratner DI et al.; To aid linkage analysis and mapping studies in Dictyostelium discoideum, we have constructed several tester strains with easily scored mutations characterizing the six currently identified linkage groups . Use has been made of conditionally lethal mutants unable to grow upon Bacillus subtilis, and the locus of the mutation involved (bsgA) has been assigned to linkage group III . The mutation cobA1, which confers resistance to cobaltous chloride, has been assigned to a previously unidentified linkage group (VII) . The temperature-sensitive growth mutation tsgC7, previously reported to define linkage group V, has been reassigned to group III, leaving linkage group V presently unmarked . The further use of genetic tester strains is described.

J Virol, 1978 Dec, 28(3), 753 - 66
Production and expression of dTMP-enriched DNA of bacteriophage SP15; Casella E et al.; Normal DNA of Bacillus subtilis phage SP15 contains approximately equimolar quantities of dTMP and a hypermodified nucleotide, 5-dihydroxypentyl-dUMP (DHPdUMP) . Deoxythymidine (dThd) rescue of phage DNA synthesis in 5-fluorodeoxyuridine (FUdR)-inhibited cultures resulted in the synthesis of SP15 DNA containing enhanced levels of dTMP and correspondingly reduced levels of DHPdUMP . This rescued system was used to probe possible roles of DHPdUMP in phage development . The results suggested that normal levels of DHPdUMP were not required for proper transcription of phage DNA, but normal amounts of DHPdUMP were indispensable for phage assembly and/or DNA maturation . The amount of exogenous dThd required to rescue phage DNA synthesis in FUdR-inhibited cultures was 20-fold higher than the concentration required to rescue cellular replication, whereas the same low concentrations of dThd sufficed to rescue viral and bacterial DNA syntheses in aminopterin-inhibited cultures . Normal SP15 DNA was made in rescued, aminopterin-inhibited cultures . We suggest that FUdR (but not aminopterin) partially suppresses biosynthesis of the hypermodified nucleotide and that there is a barrier to replacement of DHPdUMP by dTMP; therefore, exceptionally large amounts of dThd must be salvaged in FUdR-inhibited cultures to force replacement of the unusual nucleotide by dTMP.

Can J Microbiol, 1978 Dec, 24(12), 1439 - 51
The effect of chemical fixatives on cell walls of Bacillus subtilis; Beveridge TJ et al.; Cell walls of Bacillus subtilis were treated with several chemical fixatives which are commonly used preparatory to electron microscopy; i.e., osmium tetroxide, formaldehyde, acrolein, crotonaldehyde, and glutaraldehyde . Dimensional analysis was performed on thin sections of fixed walls from plastic embeddings and, by means of the statistical technique of multiple comparisons, significant differences were found between wall thicknesses from the various fixations . These differences varied with the fixation time and the type of fixative used in the reaction . When compared to embedded walls which had been stained before fixation, the overall effect was a reduction in wall thickness which was attributed to fixative action and not to the embedding or staining processes . The reduction of wall thickness was even more apparent when dimensions of fixed walls were compared to published dimensions of both frozen sections and freeze-etch profiles . Since these fixatives bind to reactive sites within the wall fabric, a change in electrochemical charge density is effected which can be monitored in terms of heavy-metal-binding capacity . Most monoaldehyde fixatives and osmium tetroxide render the wall as reactive, or less reactive, to uranyl acetate as unfixed walls, whereas glutaraldehyde can significantly increase the binding capacity.

J Gen Virol, 1978 Dec, 41(3), 563 - 72
Studies on transduction process by SPP1 phage; Ferrari E et al.; The conditions for optimal transduction efficiency of the Bacillus subtilis phage SPP1 have been investigated . By irradiating transducing lysates with u.v . light we have been able to obtain a fivefold increase in the number of transductants and to reduce strongly the interference caused by infective particles . Any dependence of SPP1 transduction on PBSX induction has been ruled out by the use of xin mutants, which are unable to induce the defective phage . SPP1 mediated transduction is susceptible to the restriction and modification system of B . subtilis . The rec functions involved in the recombination of the SPP1 transduced DNA fragment are probably identical to those required in DNA transformation and heterologous PBS1 transduction.

Biokhimiia, 1978 Dec, 43(12), 2233 - 40
{Immunochemical study of subtilisins obtained from Bacillus subtilis A-50 and some of its mutants}; Logunov AI et al.; Physico-chemical parameters of subtilisins from the original Bacillus subtilis A-50 strain (proteolytic activity, electrophoretic mobility, molecular weight, reactions with specific inhibitors) were similar to those mentioned in the literature for the enzymes of other strains . Immunological experiment has shown, that Bacillus subtilis A-50 subtilisins with various electrophoretic mobility do not differ in their antigenic properties . Enzymes with high electrophoretic mobility from mutant strains were similar to I--III subtilisin fractions from Bac . subtilis A-50 in the antigenic characteristics . However, the antigenic heterogeneity was observed in I, II and III enzyme fractions of some mutant strains . Subtilisins studied appear to form the isoenzyme system.

J Antibiot (Tokyo), 1978 Dec, 31(12), 1211 - 7
CC-1065 (NSC-298223), a new antitumor antibiotic . Production, in vitro biological activity, microbiological assays and taxonomy of the producing microorganism; Hanka LJ et al.; A new antitumor antibiotic is produced in fermentation liquors of Streptomyces zelensis sp.n . The antibiotic is biologically active at extremely low concentrations . At 40 pg/ml, it inhibited 90% of the growth of L1210 cells in culture in tube dilution assays . The minimal inhibitory concentrations against Gram-positive bacteria is between 1 approximately 10 ng/ml, while these values for Gram-negative bacteria and fungi are mostly under 1 microgram/ml . A microbiological assay with Bacillus subtilis can detect concentrations of 1 approximately 2 ng/ml.

Proc Natl Acad Sci U S A, 1978 Dec, 75(12), 5993 - 7
Thiostrepton-resistant mutants exhibit relaxed synthesis of RNA; Smith I et al.; Spontaneous mutants of Bacillus subtilis resistant to thiostrepton (TSP) exhibit relaxed synthesis of RNA when starved for required amino acids . Intact cells of tsp mutants cannot synthesize the regulatory nucleotides, ppGpp and pppGpp, after amino acid deprivation . Because ribosomes isolated from spontaneous revertants to thiostrepton sensitivity and from wild-type stringent strains can synthesize (p)ppGpp whereas ribosomes isolated from tsp strains cannot synthesize these regulatory nucleotides in the presence of stringent factor, it appears that the lesion is expressed at the level of the ribosome . Genetic mapping, via three-factor transformational crosses, has shown that tsp is closely linked to rif, in the order cysA14, tsp, rif-I, strA . The phenotype of the tsp mutants indicates that they are of the relC type . Their map position indicates that they are different from a previously described B subtilis rel mutation . Ribosomes from the latter strain can synthesize (p)ppGpp in cell-free extracts.

Proc Natl Acad Sci U S A, 1978 Dec, 75(12), 5922 - 5
Interaction of secreted nascent chains with surrounding membrane in Bacillus subtilis; Smith WP et al.; To determine the length of secreted nascent polypeptide chain that is surrounded by membrane, we digested labeled nascent chains protruding from protoplasts of Bacillus subtilis with Pronase and isolated the residual ribosome-attached chains from the membrane-polysome fraction . Gel chromatography revealed a sharp major peak that had been protected by membrane plus bound ribosomes . The ribosomes themselves protected half as great a length . Because no free chain between the ribosome and the membrane was detected by Pronase treatment, the difference between the two protected lengths should measure the length protected by the membrane . More accurate measurements of these lengths, obtained by dansylation of the exposed NH2 terminus of the isolated fragments, yielded a difference of 21 amino acids . This value corresponds to an extended chain of 75 A, which is approximately the thickness of the bacterial cell membrane . We earlier presented evidence that bacterial ribosomes are attached to membrane solely by their secreted chain . The present results further show that after loss of the extracellular segment of the chain its attachment persists, at 37 degrees as well as 0 degrees C . These findings suggest that the chain does not slip through a passive membrane but is actively held within a channel.

J Virol, 1978 Dec, 28(3), 697 - 709
Relationship of Bacillus subtilis DNA polymerase III to bacteriophage PBS2-induced DNA polymerase and to the replication of uracil-containing DNA; Hitzeman RA et al.; In vivo studies of PBS2 phage replication in a temperature-sensitive Bacillus subtilis DNA polymerase III (Pol III) mutant and a temperature-resistant revertant of this mutant have suggested the possible involvement of Pol III in PBS2 DNA synthesis . Previous results with 6-(p-hydroxyphenylazo)-uracil (HPUra), a specific inhibitor of Pol III and DNA replication in uninfected cells, suggest that Pol III is not involved in phage DNA replication, due to its resistance to this drug . Experiments were designed to examine possible explanations for this apparent contradiction . First, assays of the host Pol III and the phage-induced DNA polymerase activities in extracts indicated that a labile Pol III did not result in a labile phage-induced enzyme, suggesting that this new polymerase is not a modified HPUra-resistant form of Pol III . Indeed the purified phage-induced enzyme was resistant to the active, reduced form of HPUra under all assay conditions tested . Since in vitro Pol III was capable of replicating the uracil-containing DNA found in this phage, the sensitivity of the purified enzyme to reduced HPUra was examined using phage DNA as template-primer and dUTP as substrate; these new substrates did not affect the sensitivity of the host enzyme to the drug.

J Bacteriol, 1978 Dec, 136(3), 886 - 99
Cell wall assembly in Bacillus subtilis: location of wall material incorporated during pulsed release of phosphate limitation, its accessibility to bacteriophages and concanavalin A, and its susceptibility to turnover; Anderson AJ et al.; Addition of a pulse of phosphate to a phosphate-limited chemostat culture of Bacillus subtilis W23 led to the synthesis of teichoic acid and the consequent development by the bacteria of the ability to bind phage SP50 . In cultures growing at different rates, phage-binding properties became maximal approximately one generation time after addition of the pulse . Removal of the incorporated teichoic acid by turnover also reached its maximum rate after a similar interval . After pulsed release of phosphate limitation in B . subtilis NCTC 3610, the alpha-glucosyl residues of the incorporated teichoic acid, detected by their interaction with concanavalin A, became maximally exposed at the same time that phage binding was maximum . At that time the bacteria bound phage all over the cylindrical part of the surface and at about one-third of the polar caps . That fraction of the receptor material that is exposed soon after its incorporation was distributed along the cylindrical length of most of the bacteria, but few phages bound to the polar caps, except in the case of short bacteria; these bound phages in a markedly asymmetric manner at one pole and along their length . The significance of these results is discussed in relation to the mode of assembly of the cell wall.

J Bacteriol, 1978 Dec, 136(3), 883 - 93
Suppression of temperature-sensitive sporulation of a Bacillus subtilis elongation factor G mutant by RNA polymerase mutations; Hirochika H et al.; A class of rifampin-resistant (rfm) mutations of Bacillus subtilis suppresses the temperature-sensitive sporulation of a fusidic acid-resistant mutant . FUS426, which has an altered elongation factor G . The rfm mutation suppressed only the sporulation defect caused by the elongation factor G mutation, but could not suppress other types of induced sporulation defects . Genetic and biochemical analyses showed that the sporulation suppression by the rfm mutation was caused by a single mutation in RNA polymerase . After the early sporulation phase, the apparent rate of RNA synthesis of FUS426, measured by {3H}uracil or {3H}uridine incorporation into RNA, became lower than that of the wild-type strain, and this decrease was reversed by the rfm mutation . However, when the total rate of RNA synthesis of FUS426 was calculated by measuring the specific activity of {3H}UTP and {3H}CTP, it was higher than that of the rfm mutant, RIF122FUS426 . The possible mechanism of the functional interaction between elongation factor G and RNA polymerase during sporulation is discussed.

J Bacteriol, 1978 Dec, 136(3), 854 - 66
Alterations in Bacillus subtilis transforming DNA induced by beta-propiolactone and 1,3-propane sultone, two mutagenic and carcinogenic alkylating agents; Kubinski ZO et al.; Transforming DNA was exposed to either beta-propiolactone or 1,3-propane sultone and then used for transformation of competent bacteria to nutritional independence from tyrosine and tryptophan (linked markers) and leucine (an unlinked marker) . The ability to transform was progressively lost by the DNA during incubation with either of these two chemicals . For all three markers the inactivation curve was biphasic, with a short period of rapid inactivation followed by one characterized by a much slower rate . The overall rate of inactivation was different for all three markers and presumably was related to the size of the marker . The decrease in the transforming activity was in part due to the slower rate of penetration of alkylated DNA through the cellular membrane and its inability to enter the recipient bacteria . This decrease in the rate of cellular uptake, even for DNA eventually destined to enter the cell, began almost immediately after its exposure to the chemical and ended up with an almost complete lack of recognition of the heavily alkylated DNA by the specific surface receptors of competent cells . Such DNA attached to sites on the surface of competent bacteria which were different from receptors specific for the untreated nucleic acid . This attachment was not followed by uptake of the altered DNA . Presence of albumin during the incubation with a carcinogen further increased the degree of inactivation, indicating that the artificial nucleoproteins produced under such conditions were less efficient in the transformation assay than was the naked DNA . Cotransfomration of close markers progressively decreased, beginning immediately after the start of incubation of DNA with the chemicals . Extensively alkylated DNA fractionated by sedimentation through sucrose density gradients showed a peculiar distribution of cotransforming activity for such markers; namely, molecules larger than the bulk of DNA ("megamolecules") showed less ability to transform the second marker than did some of the apparently smaller molecules which sedimented more slowly through the gradient . An increase in cotransformation of distant markers was evident in DNA molecules after a short exposure to an alkylating agent, but cotransformation of such markers was absent in DNA treated for longer periods . The observed changes in the transforming and cotransforming activities of the alkylated DNA can be explained by what is known about the physicochemistry of such DNA and in particular about the propensity of the alkylated and broken molecules to form complexes with themselves and with other macromolecules.

J Bacteriol, 1978 Dec, 136(3), 1205 - 7
Differential effect of hydroxyurea on the replication of plasmid and chromosomal DNA in Bacillus subtilis; Shivakumar AG et al.; The replication in Bacillus subtilis of the staphylococcal R plasmids pE194, pBD15, pUB110, pSA0501, and pSA2100 has been studied in the presence of hydroxyurea . In all cases, an enrichment for covalently closed circular DNA compared with chromosomal DNA was observed . In this respect, hydroxyurea mimics the effect previously observed with pUB110, using strains carrying the conditional mutation dnaA13 . This mutation has been reported to affect ribonucleotide reductase (G . W . Bazill and D . Karamata, Mol . Gen . Genet . 117:19-29, 1972) . An explanation for these effects is offered, together with some supporting evidence.

Mol Gen Genet, 1978 Nov 29, 167(2), 157 - 64
SPP1 DNA replicative forms: growth of phage SPP1 in Bacillus subtilis mutants temperature-sensitive in DNA synthesis; Mastromei G et al.; The development of bacteriophages SPP1 and phi 29 has been studied in several B . sutilis mutants defective in host DNA replication, under non permissive conditions . Several gene products, involved in the synthesis of host DNA, are required for phi 29 replication, while SPP1 seems to require only the host DNA polymerase III . In addition both phages are unable to grow in a dna A mutant (ribonucleotide reductase) . Taking advantage of the fact that SPP1 DNA is actively replicated in several dna mutants at non-permissive temperature, we have studied the structure of the replicative intermediates of this phage in the absence of interfering host DNA synthesis . Fast sedimenting forms of SPP1 DNA can be isolated from phage infected cells and evidence of covalently joined concatemers has been obtained, suggesting the presence of terminally repeated sequences.

Biochim Biophys Acta, 1978 Nov 21, 521(1), 16 - 26
Induction of sporulation by inhibitory purines and related compounds; Heinze JE et al.; Sporulation of Bacillus subtilis can be induced, in the presence of excess ammonia, glucose and phosphate, by many purine derivatives under conditions of partial growth inhibition . Some of the compounds are known inhibitors of purine nucleotide synthesis . For most compounds the effect is counteracted by adenine and guanine . Partial growth inhibition by amethopterin (methotrexate) causes sporulation in the absence of purines but not in their presence . Unable to induce sporulation at any concentration are inhibitors of DNA, RNA, and protein synthesis as well as base or amino acid analogs that are incorporated into these polymers.

Biochim Biophys Acta, 1978 Nov 15, 544(1), 1 - 7
Regulation of lactate dehydrogenase synthesis in Bacillus subtilis; Yashphe J et al.; The regulation of lactate dehydrogenase in Bacillus subtilis was determined under a variety of growth conditions and in mutants blocked in the citric acid cycle . The synthesis of lactate dehydrogenase increased sharply concomitantly upon the exhaustion of glucose from the medium and the onset of the stationary phase . The synthesis of lactate dehydrogenase may be under catabolite repression control . Studies with mutants blocked in the citric acid cycle showed that lactate dehydrogenase is regulated independently of either the oxidative or reductase branches of the cycle . Certain citric acid cycle mutants, e.g., aconitase or succinate dehydrogenase, exhibited very low levels of lactate dehydrogenase while others, e.g., malate dehydrogenase or isocitrate dehydrogenase, showed normal levels . A stage O sporulation mutant expressed levels of lactate dehydrogenase more than one thousand-fold higher than the low group of citric acid cycle mutants . The induction of lactate dehydrogenase was shown to be independent of the accumulation of its substrate, pyruvate.

Mol Gen Genet, 1978 Nov 9, 166(3), 287 - 90
Genetic exchange in Bacillus subtilis in soil; Graham JB et al.; Genetically labelled strains of Bacillus subtilis have been shown to exchange blocks of linked genes while growing together in soil . After eight days of incubation, 79% of unselected colony-forming units exhibited a phenotype containing markers from both parents; the parental strains were not detected after the first day of incubation . High frequencies of transformation were also obtained by adding genetically labelled deoxyribonucleic acid to single-strain soil cultures . Observed linkage of genetic markers was greater in soil transformation than in standard laboratory procedures . The results indicate that transformation may play an important role in the adaptation of the Bacilli to their natural habitat.

Ann Microbiol (Paris), 1978 Nov-Dec, 129 B(4), 537 - 49
Pleiotropic control mutations affecting the sporulation of Bacillus subtilis; Balassa G et al.; Mutations affecting quantitatively the production of the sporulation-associated extracellular alkaline protease were isolated and characterized . They fall into at least five genes, three of which, ScoA, B and C, were mapped in the argC-metC region . The pleiotropic effects of these mutations concern several or all of the following: rate and timing of protease production, synthesis of alkaline phosphatase, time-course of spore formation . Electron microscopic evidence indicates delayed switch from one morphological stage to another . The nature of the Sco mutations and the genetic regulation of sporulation are discussed.

Prikl Biokhim Mikrobiol, 1978 Nov-Dec, 14(6), 890 - 7
{Preparation of extracellular ribonuclease from Bacillus subtilis}; Bashkis EV et al.; The paper describes a method for isolating alkaline ribonuclease from the culture liquid Bacillus subtilis KP 349 which involved: submerged cultivation of the producer on complex and synthetic nutrient media with optimized RNase activity, acid treatment of the total culture liquid, and filtration through perlite . Further treatment may include either spray drying of the culture liquid filtrate or its concentration in a vacuum evaporator, dialysis of the concentrate against distilled water, and dialyzate lyophilization . As a result, commercial RNase preparations with activities of 30--60 thous . and 160--300 thous . units/g, respectively, were obtained . The enzyme purification was carried out by chromatography and rechromatography on phosphocellulose columns . The resultant RNase of Bac . subtilis had a specific activity of 41--44 thous . units/mg protein, contained no nonspecific phosphodiesterase, DNase, acid or alkaline phosphomonoesterases.

Can J Microbiol, 1978 Nov, 24(11), 1431 - 3
Survival of microbial films in the microwave oven; Page WJ et al.; Air-dried films of Escherichia coli, Saccharomyces cerevisiae, and Bacillus subtilis spores on membrane filters, exposed to 10 min full power (650 W, 2450 MHz) irradiation in a Toshiba model ER776BT microwave oven, showed a 5-, 2-, and 0-log reduction of viable organisms respectively . Suspensions of cells or spores in phosphate buffer treated under similar conditions showed 8 logs of killing within 30 s (S . cerevisiae), 45 s(E . coli), and by 10 min (B . subtilis spores) of exposure.

Genetika, 1978 Nov, 14(11), 2046 - 8
{UV-light induction of "true" revertants to adenine independence in Bacillus subtilis cells}; Lotareva OV; The UV-irradiation of Bacillus subtilis Mu5u8u16 (met5 leu8 purA) induces with relatively high frequency the revertants to adenine independence (Ade+) which form the rapidly growing morphologically uniform colonies on the solid selective medium . The genetic analysis of a portion of UV-induced Ade+ revertants (crosses in transformation system) has cleared up that their DNA does not contain the original mutation ade16 . This means that they arise as the result of "true" reversions . This reversion in purA gene can serve as a good model for the study of UV-induced mutagenesis in a proximal structural locus of Bac . subtilis chromosome.

Genetika, 1978 Nov, 14(11), 1900 - 7
{Effect of Bacillus subtilis A-50 "streptomycin" mutations on the formation of molecular forms of subtilisin, an extracellular alkaline proteinase}; Ermakova LM et al.; The patterns of subtilisin molecular forms of streptomycin-resistant (Strr) and streptomycin-dependent (Strd) mutants of Bacillus subtilis A-50, as well as the revertants of Strd to streptomycin-independence (Str1) were studied . Strr mutants had different quantitative pattern of the same subtilisin molecular forms as compared with the initial strain A-50 (the forms with Rf 0.08, 0.16 and 0.3) . In comparison with the initial strain A-50, Strd mutants and Str1 revertants revealed three additional forms of the active enzyme with Rf 0.02, 0.5 and 0.7 and the molecular weights less than 35,000, 28,000 and 20,000 respectively . It was suggested that the rate and character of the enzyme secretion of the degree of its post-translational modifications might result in the different pattern of subtilisin molecular forms produced by these streptomycin mutants.

Biull Eksp Biol Med, 1978 Nov, 86(11), 556 - 7
{Effect of dimethyl sulfoxide on transformation of B . subtilis in vitro}; Orlova EB et al.; Bacillus subtilis transformation was conducted in the presence of dimethylsulfoxide and polyethyleneglycol . B . subtilis transformation was most frequent under the effect of 0.1% dimethylsulfoxide and was not altered significantly by polyethyleneglycol . As suggested, an increase of the transformation frequency was associated with the change of the membrane permeability under the influence of dimethylsulfoxide of with the altered activity of the membrane DNase.

J Bacteriol, 1978 Nov, 136(2), 818 - 21
Tunicamycin-resistant mutants and chromosomal locations of mutational sites in Bacillus subtilis; Nomura S et al.; The types of tunicamycin-resistant mutants of Bacillus subtilis were analyzed, and their mutational sites on the chromosome were mapped . A type 1 mutation that simultaneously expressed hyperproductivity of extracellular alpha-amylase was located close to amy E . Type 2 mutations were near aroI.

J Bacteriol, 1978 Nov, 136(2), 799 - 802
Chromosome-membrane association in Bacillus subtilis . IV . Further purification of DNA-membrane complex by using a combination of centrifugation and electrophoresis; Toyoda H et al.; We have developed a simple procedure to purify a DNA-membrane complex from Bacillus subtilis by using a combination of centrifugation and electrophoresis . Several unique proteins were detected in the purified complex.

J Bacteriol, 1978 Nov, 136(2), 484 - 90
Inhibition of Bacillus subtilis spore germination by various hydrophobic compounds: demonstration of hydrophobic character of the L-alanine receptor site; Yasuda-Yasaki Y et al.; L-Alanine-initiated germination of Bacillus subtilis spores was inhibited by various kinds of hydrophobic compounds . Good correlation of inhibitory effect with hydrophobicity of the compound was demonstrated by using regression analysis in which the hydrophobic character was expressed by the partition coefficient in an octyl alcohol-water system . The correlation coefficient for 20 alcohols was 0.959, and that for 19 miscellaneous compounds was 0.906 . Regression lines of the alcohols and other hydrophobic compounds were almost identical, showing that hydrophobic interaction played an important role in inhibition . Diphenylamine was one of the most effective inhibitors examined . n-Octyl, n-nonyl, and n-decyl alcohols were the most effective alcohols . The mode of inhibition by diphenylamine and n-octyl alcohol was a "mixed type" (competitive plus noncompetitive type) with respect to L-alanine; that by D-alanine was competitive inhibition . Sites for diphenylamine, n-octyl alcohol, and D-alanine may have overlapped . Inhibition was reversible by washing; heat resistance, stainability, and germination rate of the washed spores remained unaltered . Thus, we confirmed that the inhibition may occur before the initial trigger reaction of germination and that it may be due to the interaction between a hydrophobic compound and a hydrophobic region closely associated with the L-alanine receptor site on the spore.

Cancer Res, 1978 Nov, 38(11 Pt 1), 3918 - 21
Carcinogenicity of triethanolamine in mice and its mutagenicity after reaction with sodium nitrite in bacteria; Hoshino H et al.; Mice fed a diet containing 0.3 or 0.03% triethanolamine developed malignant tumors . Females showed a high incidence of tumors in lymphoid tissues, while this type was absent in males . Tumors in other tissues were produced at a considerable rate in both sexes, but no hepatoma was found . Triethanolamine was not mutagenic to Bacillus subtilis by itself, but it became mutagenic after reacting with sodium nitrite under acidic conditions or when the mixture was heated . Although N-nitrosodiethanolamine, a known carcinogen and mutagen, was detected in the reaction mixture by thin-layer chromatography, it may not be the main mutagenic product, because the product was a stable and direct mutagen and its mutagenic activity was destroyed by liver enzymes, unlike N-nitrosodiethanolamine . The lethal and mutagenic DNA damages produced by this unidentified product were susceptible to some extent to the repair functions of the bacteria.

J Bacteriol, 1978 Nov, 136(2), 625 - 30
Benzeneboronic acid selectively inhibits sporulation of Bacillis subtilis; Davis-Mancini K et al.; m-Aminobenzeneboronic acid at levels of 0.2 mM in nutrient broth medium selectively inhibited sporulation without appreciably altering vegetative growth . Significant inhibitory effects were seen even when it was added as late as 6 h after the end of logarithmic growth . The pH changes associated with growth and sporulation of Bacillus subtilis in nutrient broth were not significantly altered by the inhibitor . When it was present in cultures of actively growing cells, its inhibitory effect could not be reversed by simple dilution . The compound caused extensive clumping, of cells, which appeared not to be related to the ability of boronates to esterify to diols.

Mol Gen Genet, 1978 Oct 30, 166(2), 119 - 26
An unstable donor-recipient DNA complex in transformation of Bacillus subtilis; Popowski J et al.; In re-extracted DNA obtained shortly after uptake of transforming DNA by Bacillus subtilis, increased amounts of donor DNA radioactivity banding at the position of donor-recipient DNA complex (DRC) are observed in CsCl gradients, if the cells are irradiated with high doses of UV prior to reextraction of the DNA . Qualitatively, the same phenomenon is observed if lysates of transforming cells are irradiated . UV-irradiation of lysates of competent cells to which single-stranded DNA is added after lysis, does not result in linkage of this DNA to the chromosomal DNA . Two observations argue in favour of the formation of a specific labile complex between donor and resident DNA during transformation . Firstly, heterologous donor DNA from Escherichia coli, although being processed to single-stranded DNA in competent B . subtilis, does not seem to be linked to the recipient chromosome upon UV-irradiation, and secondly, the labile complex of donor and recipient DNA can be stabilized by means of treatment of the lysates of transforming cells with 4, 5(1), 8-trimethylpsoralen in conjuction with long-wave ultra violet light irradiation . This indicates that basepairing is involved in the formation of the complex . On the basis of these results we assume that the unstable complex of donor and recipient DNA is an early intermediate in genetic recombination during transformation.

Eur J Biochem, 1978 Oct 16, 90(3), 581 - 3
Restriction and modification in Bacillus subtilis . Localization of the methylated nucleotide in the BsuRI recognition sequence; Gunthert U et al.; Calf thymus DNA was methylated in vitro with cell extracts of Bacillus subtilis OG3R (r+m+) and S-adenosyl{Me-3H}methionine . After depurination of the {3H}methylated DNA, the analysis of the pyrimidine dinucleotides revealed the following positions of the methylated nucleosides (indicated by an asterisk) within the BsuRI recognition sequence: 5' dG--dG--dC--dC dC--dC--dG--dG 5'.

J Biol Chem, 1978 Oct 10, 253(19), 6694 - 701
Spore coat protein of Bacillus subtilis . Structure and precursor synthesis; Munoz L et al.; The coat protein of Bacillus subtilis spores comprises about 10% of the total dry weight of spores and 25% of the total spore protein . One protein with a molecular weight of 13,000 to 15,000 comprises a major portion of the spore coat . This mature spore coat protein has histidine at its NH2 terminus and is relatively rich in hydrophobic amino acids . Netropsin, and antibiotic which binds to A-T-rich regions of DNA and inhibits sporulation, but not growth, decreased the synthesis of this spore coat protein by 75% . A precursor spore coat protein with a molecular weight of 25,000 is made initially at t1 of sporulation and is converted to the mature spore coat protein with a molecular weight of 13,500 at t2 - t3 . These data indicate that the spore coat protein gene is expressed very early in sporulation prior to the modifications of RNA polymerase which have been noted.

Biochim Biophys Acta, 1978 Oct 4, 512(3), 489 - 94
The influence of branched-chain and omega-alicyclic fatty acids on the transition temperature of bacillus subtilis lipids; Blume A et al.; The influence of branched-chain and omega-alicyclic fatty acids on the transition temperature of Bacillus subtilis lipids was studied by measuring the fluorescence depolarisation of the probe 1,6-diphenyl-1,3,5-hexatriene incorporated into lipid bilayers . Only anteiso-C15 and C17 fatty acid-enriched lipids showed no transition in the observed temperature range . Compared to the transition of normal lipids iso-fatty acid-enriched lipids have a slightly higher transition temperature . The incorporation of omega-alicyclic fatty acids with increasing size of the alicycle leads to a decrease in the transition temperature . A possible role of omega-cyclohexane fatty acids in Bacillus acidocaldarius is proposed.

Ann Microbiol (Paris), 1978 Oct, 129 B(3), 377 - 90
Ultrastructural effects of the chemical agents and moist heat on bacillus subtilis . II.--Effects on sporulating cells; Sousa JC et al.; When sporulating cells are treated with the organic solvents (xylene, toluene, octanol and chloroform), with TCA or with moist heat (10 min at 80 degree C), the sporangial cells exhibit the same ultrastructural changes as do the vegetative cells . The forespores undergo similar changes after early but not after late treatment . Resistance toward the killing effect and toward ultrastructural alterations appear in the same order . One could correlate the appearance of resistances with precise ultrastructural events as follows:--xylene resistance: cortex formation (stage IV);--resistance to toluene, octanol and chloroform: coat development and cortex maturation (respectively early, middle and late stage V);--TCA and heat resistance: spore maturation (stage VI) . The possible mechanisms of the chemical resistances are discussed.

Ann Microbiol (Paris), 1978 Oct, 129 B(3), 363 - 75
Ultrastructural effects of chemical agents and moist heat on Bacillus subtilis . I--Effects on vegetative cells; Silva MT et al.; The ultrastructural alterations induced by treatment of vegetative Bacillus subtilis cells with organic solvents, trichloroacetic acid (TCA) and moist heat were examined by electron microscopy . Organic solvents disorganize the membrane and change the asymmetric unit membrane profile to a symmetric profile . They also lead to partial solubilization of the membrane and produce small or extensive gaps . Membrane damaging activity increases in the following order: xylene = toluene less than octanol less than choroform . TCA coagulates the cytoplasm which shows large, electron-dense, pronase-sensitive blocks . Moist heat alters both the membrane and the cytoplasm.

Ann Microbiol (Paris), 1978 Oct, 129 B(3), 339 - 62
Ultrastructure and development of an exosporium-like outer spore envelope in Bacillus subtilis; Sousa JC et al.; An exosporium-like outermost envelope is occasionally observed in thin sections of Bacillus subtilis spores . Treatment of the mature spores with urea and mercaptoethanol (sometimes completed by sodium dodecyl sulfate) or with NaOH, disorganizes and partially solubilizes the outer spore coat . This treatment permits a clear visualization of the exosporium in all spores of several B . subtilis strains . Exosporium appears either as a single sheet, 8-9 nm thick, or with a triple-layered unit membrane-like profile . Frequently it exhibits a crystal-like periodic pattern . The exosporium primordium appears first at stage IV, and its development is apparently independent of the formation of the cortex and of the spore coats . No morphological relationship was found between the outer forespore membrane and the exosporium.

Biochem Genet, 1978 Oct, 16(9-10), 867 - 81
An anthranilate synthase of the extreme aminase type in a species of blue-green bacteria (algae); Friedman E et al.; Anthranilate synthase of Agmenellum quadruplicatum, a unicellular species of blue-green bacteria, consists of two nonidentical subunits . A 72,000 dalton protein has aminase activity but is incapable of reaction with glutamine (amidotransferase) unless a second protein (18,000 molecular weight) is present . The small subunit was first detected through its ability to complement a partially purified aminase subunit from Bacillus subtilis to produce a hybrid complex capable of amidotransferase function . Conditions for the function of the heterologous complex were less stringent than for the homologous A . quadruplicatum complex . A reducing agent such as dithiothreitol stabilizes the A . quadruplicatum aminase subunit and is obligatory for amidotransferase function . L-Tryptophan feedback inhibits both the aminase and amidotransferase reactions of anthranilate synthase; Ki values of 6 X 10(-8) M for the amidotransferase activity and 2 X 10(-6) M for the aminase activity were obtained . The Km value calculated for ammonia (2.2 mM) was more favorable than the Km value glutamine (13 mM) . Likewise, the Vmax of anthranilate synthase was greater with ammonia than with glutamine . Starvation of a tryptophan auxotroph results in a threefold derepression of the aminase subunit, but no corresponding increase in the small 18,000 M subunit occurs . While microbial anthranilate synthase complexes are remarkably similar overall, the relatively good aminase activity of the A . quadruplicatum enzyme may be of physiological significance in nature.

Mutat Res, 1978 Oct, 52(1), 49 - 56
Study of MFD in Bacillus subtilis; Filippov VD et al.; The frequency of UV-induced extragenic suppressor reversions to leucine independence in B . subtilis carrying a leu8 mutation decreased when irradiated cells were temporarily incubated in medium deprived of nitrogen sources . This mutation frequency decline (MFD) was inhibited by acriflavine and was poorly expressed in a uvr1 mutant . Consequently, MFD may be considered as the manifestation of an anti-mutagenic activity of excision repair . MFD was decelerated and even vanished in cells subjected to prolonged starvation of nitrogen sources before irradiation . MFD was accelerated in bacteria that were first irradiated and incubated in nutritional medium for at least 30 min . The stimulation of MFD by UV exposure was observed only in the uvr+ strain and depended on protein synthesis after irradiation . It is assumed that different rates of MFD in cells of various pre-radiation histories reflect different levels of the excision-repair activity inherent in these cells.

Genetika, 1978 Oct, 14(10), 1696 - 1705
{Amplification of the riboflavin operon genes of Bacillus subtilis in Escherichia coli cells}; Rabinovich PM et al.; Amplification of Bacillus subtilis DNA fragments was performed in Escherichia coli using plasmid RSF2124 . The main principle of isolation and cloning hybrid plasmids was described using genes of riboflavin operon as a model . Bac . subtilis DNA was treated with restriction endonuclease EcoR; followed by the agarose gel electrophoretic separation of the resulting fragments . Gels were sliced, DNA was eluted from the corresponding slices and used to transform Bac . subtilis auxotrophs rib A72, rib S110 and rib D107 . DNA fraction with the molecular weight 7 . 10(6) daltons restored prototrophy of these mutants . DNA of this fraction was ligated with EcoRI treated plasmid RSF2124 DNA and used for transformation of E . coli rk-mk+ . Ampicillin resistant transformants which had lost the colicin production ability, were selected . The presence of riboflavin genes within the hybrid plasmids was detected by transformation of B . subtilis auxotrophs . Three hybrid plasmids (pPR1, pPR2 and pPR3), containing a fragment of Bac . subtilis DNA with the molecular weight 6.8 . 10(-6) daltons including riboflavin operon, were selected . The analysis of the transformation activity of Bac . subtilis DNA and plasmid pPR1 DNA revealed, that there was no restriction activity of Bac . subtilis cells against plasmid DNA amplified in E . coli . Heteroduplex analysis has shown that plasmids pPR1 and pPR2 differ in the orientation of Bac . subtilis DNA fragment . DNA of these plasmids restored prototrophy of the several studied E . coli riboflavin auxotrophs.

J Bacteriol, 1978 Oct, 136(1), 304 - 11
Characterization of succinic dehydrogenase mutants of Bacillus subtilis by crossed immunoelectrophoresis; Rutberg B et al.; Eleven succinic dehydrogenase (SDH) mutants in Bacillus subtilis were analyzed by crossed immunoelectrophoresis with antiserum prepared against wild-type B . subtilis cytoplasmic membrane . A precipitate which stained for SDH was found in Triton X-100-solubilized wild-type membranes and in membranes from two of the SDH mutants . The remaining nine mutants did not show an SDH-staining precipitate . The respective mutations in these nine mutants all map in one locus, citF (Ohne et al., J . Bacteriol . 115:738-745, 1973) . An SDH-specific antiserum was prepared by immunizing rabbits with the SDH precipitate obtained in crossed immunoelectrophoresis with solubilized wild-type membrane . Using this antiserum, it was shown that all of the nine citF mutants lack an SDH-specific antigen in the membrane but five of the citF mutants have a soluble SDH-specific antigen . No major differences were found in sodium dodecyl sulfatepolyacrylamide gels of membrane proteins from wild-type B . subtilis and from SDH mutants . A model for the organization of SDH in B . subtilis is proposed.

J Bacteriol, 1978 Oct, 136(1), 111 - 6
Independence of Bacillus subtilis spore outgrowth from DNA synthesis; Ginsberg D et al.; The outgrowth of spores of Bacillus subtilis 168 proceeded normally in temperature-sensitive DNA mutants under restrictive conditions and in the absence of DNA synthesis . Two inhibitors of DNA synthesis, nalidoxic acid and 6-(p-hydroxyphenylazo)-uracil, inhibited spore outgrowth under some nutritional conditions; this inhibition of outgrowth however, though not that of DNA synthesis, could be reversed by glucose . The sensitivity of the outgrowing spores to nalidixic acid and 6-(p-hydroxyphenylazo)-uracil inhbition decreased as a function of outgrowth time . The cells became completely resistant to the inhibitors after 90 min . The development of this resistance occurred also in the absence of DNA synthesis . It was concluded that DNA synthesis is not needed for spore outgrowth, and that outgrowing cells and vegetative cells differ in their sensitivity to these inhibitors.

J Bacteriol, 1978 Oct, 136(1), 10 - 8
Detergent-resistant variants of Bacillus subtilis with reduced cell diameter; Tilby MJ; Variants of Bacillus subtilis resistant to the detergent Triton X-100 may exhibit: (i) normal cell morphology, (ii) reduced cell diameter, or (iii) helical cell shape . One variant of type ii was studied in some detail . Triton resistance, cell diameter reduction, and poor sporulation all may have resulted from a single mutation . High concentrations of Triton caused rapid lysis of wild-type cells . B . subtilis adapted to low Triton concentrations such that, upon subsequent exposure to higher concentrations, growth continued, although it bacame inhibited at very high concentrations . The variant studied retained its sensitivity to Triton-induced lysis but, after adaptation, grew at very high Triton levels . In this strain, cell diameter and cross-sectional area were reduced to about 73 and 50%, respectively, of those of wild type, yet the cells grew at normal rates, and DNA/protein/RNA ratios were largely unaltered . Peptidoglycan content per unit of cell surface area was higher in the variant than in the wild type under at least certain growth conditions.

J Virol, 1978 Oct, 28(1), 403 - 7
Restriction endonuclease mapping of bacteriophage phi105 and closely related temperate Bacillus subtilis bacteriophages rho10 and rho14; Perkins JB et al.; Cleavage maps of the three similar Bacillus subtilis temperate bacteriophages, phi105, rho10, and rho14, were constructed by partial digestion analysis utilizing the restriction endonuclease EcoRI . Comparison of the topography of these maps indicates that all phage DNAs posses cohesive ends and a number of EcoRI restriction sites; the fragments are conserved, and the estimated base substitution/nucleotide divergence between these phages is 0.03 to 0.07 based on conserved fragments or between 0.03 and 0.11 based on conserved cleavage sites . These lines of evidence indicate that phi105, rho10, and rho14 are closely related . Double-enzyme digestion analysis reveals that rho14 DNA has unique SalGI and BglII restriction sites and phi105 DNA has a unique SalGI restriction site, making these phages possible cloning vectors for B . subtilis.

J Virol, 1978 Oct, 28(1), 395 - 402
Correlated genetic and EcoRI cleavage map of Bacillus subtilis bacteriophage phi105 DNA; Scher BM et al.; The seven previously identified EcoRI cleavage fragments of phi 105 DNA were ordered with respect to their sites of origin on the phage genome by marker rescue . One fragment, H, did not carry any determinants essential for replication . This fragment was totally missing in a deletion mutant which exhibited a lysogenization-defective phenotype . There is a nonessential region on the phi 105 genome which begins in fragment B, spans fragment H, and ends in fragment F . The size of the nonessential region, as estimated by alterations observed in the fragmentation patterns of deletion mutant DNAs, is approximately 2.7 X 10(6) daltons . Two new EcoRI cleavage fragments with molecular weights of approximately 0.2 X 10(6) were detected by autoradiography of 32P-labeled DNA . These small fragments were not located on the cleavage map.

Biochemistry, 1978 Sep 19, 17(19), 3992 - 6
Structure of iturine A, a peptidolipid antibiotic from Bacillus subtilis; Peypoux F et al.; A mixture of iturines extracted from Bacillus subtilis gave, on column chromatography, iturine A, iturine B, and iturine C . Iturine A has the entire antifungal activity . It is a mixture of two homologous peptidolipids C48,H74N12O14 and C49H76N12O14 (mp 177 degrees C, {alpha}D-1.7 degrees in methanol (c 0.05 g/mL); mol wt 1042 and 1056) . The lipid moiety is a mixture of 3-amino-12-methyltridecanoic acid and 3-amino-12-methyltetradecanoic acid . The peptide moiety contains 7 mol of amino acids: D-Asn2, L-Asn, L-Gln, L-Pro, L-Ser, and D-Tyr . A cyclic structure for iturine A with the serine residue linked to the fatty amino acids through a peptide bond has been domonstrated . By mild HCl hydrolysis, lipid-soluble and water-soluble peptides were obtained . They were analyzed by chemical methods and by mass spectrometry . Permethylated and perdeuteriomethylated derivatives of iturine A were also subjected to mass spectrometric analysis . Both chemical analysis and mass spectrometry led to the cyclic structure I for iturine A.

Biochem J, 1978 Sep 15, 174(3), 1079 - 82
Interference by branched-chain amino acids in the assay of alanine with alanine dehydrogenase; Lund P et al.; Commercial preparations of alanine dehydrogenase from Bacillus subtilis are contaminated to varying extents with activity towards branched-chain amino acids . The Km values for these amino acids are of the same order as for L-alanine (about 10(-3)M) . The branched-chain amino acid dehydrogenase activity is lost on dialysis for 2--4h against water or 2mM-EDTA.

Experientia . 1978 Sep 15;34(9):1158.
Porphobilinogen-accumulation by a porphyrin auxotrophic strain of Bacillus subtilis; Miczak A et al.; The amount of porphobilinogen accumulated by a Bacillus subtilis hemC mutant strain is dependent on time and exogenic aminolevulinic acid addition . hemC mutant seems to be suitable for porphobilinogen production on a large scale.

Mol Gen Genet, 1978 Sep 8, 164(3), 335 - 9
Bacillus subtilis dnaF: a mutation of the gene specifying the structure of DNA polymerase III; Vrooman MJ et al.; The characteristics of Bacillus subtilis dnaF, a mutation specifying a temperature sensitive phenotype, were examined to determine its relationship to polC, the gene specifying the structure of DNA polymerase III (pol III) . Exposure of growing cells bearing dnaF to non-permissive temperature inhibited replicative DNA synthesis and specifically depressed the expression of pol III activity in crude extracts . Highly purified pol III derived from cells bearing dnaF was temperature.sensitive in its polymerase activity, indicating that dnaF is a specific, polC mutation which specifies a structurally altered enzyme.

Genetika, 1978 Sep, 14(9), 1530 - 8
{Riboflavin biosynthesis operon of Bacillus subtilis . XIV . Operator-constitutive mutants}; Bresler SE et al.; Numerous operator-constitutive mutants of riboflavin biosynthesis were selected . All of them map in a short region of the Bacillus subtilis chromosome . The yield of riboflavin synthetase from this mutant is different, but in most cases much lower than the maximal yield from a repressor minus strain . Our tentative explanation is a partial overlap of the sites for the adsorption of repressor and RNA-polymerase . Therefore the affinity to the transcribing enzyme is diminished in the operator constitutive strains . The affinity of the repressor-effector complex to the operator depends on the effector structure.

Biokhimiia, 1978 Sep, 43(9), 1539 - 48
{The role of a protonmotive force in genetic transformation of Bacillus subtilis}; Griniuvene BB et al.; The hypothesis on the role of protonmotive force in the transport of DNA through the membrane of Bac . subtilis cell during initial stages of genetic transformation was tested . A genetic transformation of arsenate-treated cells was observed . Treatment of cells by the protonophorous uncoupler of oxidative phosphorylation-carbonylcyanide dichlorophenyl--hydrazone-led to the inhibition of initial stages of genetic transformation having no significant effect on the level of intracellular ATP concentration and on the viability of cells . The dissipation of protonmotive force by means of K+ and H+ fluxes catalyzed by valinomycin and nigericin also caused the inhibition of initial stages of genetic transformation . The inhibitory effect of cationic penetrant tetraphenyl phosphonium was observed, the effect being potentiated by low concentrations of anionic penetrant phenyldicarbaundecaborate . The value of the membrane potential in the energized valinomycin-treated cells calculated from the distribution of K+ was within the range of 70--100 mV (inside minus) . These results support the conception that a protonmotive force drives DNA transport through the membrane of Bac . subtilis cells.

Mikrobiologiia, 1978 Sep-Oct, 47(5), 893 - 9
{Physiological and biochemical characteristics of Bacillus subtilis variants that form amylase}; Bakhmatova IV et al.; Physiologo-biochemical properties of four Bacillus subtilis variants R-623 (R, S, P and M) were studied; the variants were isolated by inoculation of the cell population . All the four variants were shown to belong to the studied species and to possess common and specific properties . They differed in 11 characters: the size of cells and spores, the ability to hydrolyse starch, the mode of growth on specific media, the spectrum of antibiotic action, etc . Three variants (R, S and P) produced alpha-amylase, the M variant did not produce the enzyme.

Eur J Biochem, 1978 Sep 1, 89(2), 389 - 95
The quaternary structure of the sheaths of defective phages similar to PBS X; Cremers AF et al.; The contractile sheaths of five defective, PBS X-like bacteriophages from Bacillus subtilis and B . licheniformis were investigated by electron microscopy, dodecylsulphate gel electrophoresis and immunodiffusion . Electron microscope images of the extended and contracted sheaths were of similar appearance, although their lengths were different . The surface lattices of both the extended and the contracted sheaths were determined by optical diffraction . This showed that the quaternary structure of the sheaths of all five defective phages originated from identical surface lattices, which could be approximately expressed by the selection rules L = -2n' + 3m and L = 9N' + 17M for the extended and contracted sheaths respectively, in which 6n' = n with n = 0 or an integer multiple of 6 . These results indicated that the packing of the protein subunits in these sheaths differed from those of other bacteriophages, for example T4 and millimicron {Amos and Klug, J . Mol . Biol . 99, 51--73 (1975); Admiraal and Mellema, J . Ultrastruct . Res . 56, 48--64 (1976)} . The molecular weight of the main sheath protein of the defective phages, as determined by dodecylsulphate gel electrophoresis, was approximately 50000 . This value differed from that for T4, but was similar to that of millimicron {Admiraal and Mellema, J . Ultrastruct . Res . 56, 48--64 (1976); King and Laemmli, J . Mol . Biol, 75, 315--337 (1973)} . The results of immunodiffusion experiments, however, pointed to a chemical difference between the sheath proteins of the defective phages and millimicron, in addition to T4.

J Virol, 1978 Sep, 27(3), 831 - 4
Evidence that the neck appendages are adsorption organelles in Bacillus subtilis bacteriophage phi29; Moreno F et al.; A mutant of Bacillus subtilis unable to adsorb phage phi29 efficiently has been isolated . This mutant can be infected by host range mutants of the phage . Since the host range mutations map in cistron 12, which codes for neck appendage protein, this would tend to confirm that these organelles are involved in viral adsorption.

J Virol, 1978 Sep, 27(3), 776 - 83
Genome-linked protein associated with the 5' termini of bacteriophage phi29 DNA; Yehle CO; A DNA-protein complex was isolated from Bacillus subtilis bacteriophage phi29 by sucrose gradient sedimentation or gel filtration in the presence of agents known to break noncovalent bonds . A 28,000-dalton protein was released from this complex by subsequent hydrolysis of the DNA . The DNA-protein complex was examined for its susceptibility to enzymes which act upon the 5' and 3' termini of DNA molecules . It was susceptible to exonucleolytic degradation from the 3' termini by exonuclease III but not from the 5' termini by lambda exonuclease . Attempts to label radioactively the 5' termini by phosphorylation with T4 polynucleotide kinase were unsuccessful despite prior treatment with alkaline phosphatase or phosphatase treatment of denatured DNA . Removal of the majority of the bound protein by proteolytic digestion did not increase susceptibility . These results suggest that the linked protein is covalently attached to the 5' termini of phi29 DNA.

J Virol, 1978 Sep, 27(3), 725 - 37
Bacillus subtilis bacteriophages SP82, SPO1, and phie: a comparison of DNAs and of peptides synthesized during infection; Lawrie JM et al.; The genomes of Bacillus subtilis phages phie, SPO1, and SP82 were compared by DNA-DNA hybridization, analysis of DNA fragments produced by digestion with restriction endonucleases, comparison of the arrays of peptides synthesized during infection, and phage neutralization . DNA-DNA hybridization experiments indicated that about 78% of the SP82 DNA was homologous with SPO1 DNA, whereas 40% of the phie DNA was homologous to either SPO1 or SP82 DNA . Agarose gel electrophoresis was used to compare the molecular weights of DNA fragments produced by cleavage of SP82, SPO1, and phie DNAs with the restriction endonucleases Hae III, Sal I, Hpa II, and Hha I . Digestion of the DNAs with Hae III and Sal I produced only a few fragments, whereas digestion with Hpa II and Hha I yielded 29 to 40 fragments, depending on the DNA and the enzyme . Comparing the Hpa II fragments, 51% of the SP82 fragments had mobilities which matched those of SPO1 fragments, 32% of the SP82 fragments matched the phie fragments, and 34% of the SPO1 fragments matched the phie fragments . Comparing the Hha I digestion products, 62% of the SP82 fragments had mobilities matching the SPO1 fragments, 24% of the SP82 fragments matched the phie fragments, and 22% of the SPO1 fragments matched the phie fragments . Analysis of peptides by electrophoresis on one-dimensional sodium dodecyl sulfate-polyacrylamide slab gels showed that approximately 70 phage-specific peptides were synthesized in the first 24 min of each infection . With mobility and the intervals of synthesis as criteria, 66% of the different SP82 peptides matched the SPO1 peptides, 34% of the SP82 peptides matched the phie peptides, and 37% of the SPO1 peptides matched the phie peptides . Phage neutralization assays using antiserum to SP82 yielded K values of 510 for SP82, 240 for SPO1, and 120 for phie.

J Bacteriol, 1978 Sep, 135(3), 952 - 60
Spore coat protein synthesis in cell-free systems from sporulating cells of Bacillus subtilis; Nakayama T et al.; Cell-free systems for protein synthesis were prepared from Bacillus subtilis 168 cells at several stages of sporulation . Immunological methods were used to determine whether spore coat protein could be synthesized in the cell-free systems prepared from sporulating cells . Spore coat protein synthesis first occurred in extracts from stage t2 cells . The proportion of spore coat protein to total proteins synthesized in the cell-free systems was 2.4 and 3.9% at stages t2 and t4, respectively . The sodium dodecyl sulfate-urea-polyacrylamide gel electrophoresis patterns of immunoprecipitates from the cell-free systems showed the complete synthesis of an apparent spore coat protein precursor (molecular weight, 25,000) . A polypeptide of this weight was previously identified in studies in vivo (L.E . Munoz, Y . Sadaie, and R.H . Doi, J . Biol . Chem., in press) . The synthesis in vitro of polysome-associated nascent spore coat polypeptides with varying molecular weights up to 23,000 was also detected . These results indicate that the spore coat protein may be synthesized as a precursor protein . The removal of proteases in the crude extracts by treatment with hemoglobin-Sepharose affinity techniques may be preventing the conversion of the large 25,000-dalton precursor to the 12,500-dalton mature spore coat protein.

J Bacteriol, 1978 Sep, 135(3), 731 - 40
Early stages in Bacillus subtilis transformation: association between homologous DNA and surface structures; Garcia E et al.; The addition of ethylenediaminetetraacetate to competent cultures of Bacillus subtilis irreversibly inhibited the transformability as well as the cellular binding of DNA . Our results show that the inhibition of DNA binding by ethylenediaminetetraacetate in whole cells, protoplasts, and membrane vesicles is mainly due to a permanent alteration of the DNA receptors . Transformation absolutely requires free magnesium ions, whereas DNA binding is a magnesium-independent step . In contrast to ethylenediaminetetraacetate, the absence of Mg2+ does not irreversibly affect the capacity of the competent cells to be transformed DNA-binding receptors located at the cell surface remain associated with the plasma membrane after protoplasting and after isolation of membrane vesicles . A Mg2+-dependent endonucleolytic activity associated with the membrane appears to be responsible for the lower levels of binding by protoplasts in the presence of this ion.

J Bacteriol, 1978 Sep, 135(3), 1158 - 61
DNA synthesis in competent Bacillus subtilis cells; Loveday KS; Competent cells of Bacillus subtilis incorporate degradation products from transfecting DNA into their chromosomal DNA . The sensitivity of this incorporation to inhibitors of bacterial DNA synthesis {phage infection or 6-(p-hydroxyphenylazo)-uracil} suggests that semiconservative DNA synthesis can occur in competent cells.

J Bacteriol, 1978 Sep, 135(3), 1149 - 50
Kasugamycin-resistant mutants of Bacillus subtilis; Tominaga A et al.; Kasugamycin-resistant mutants of Bacillus subtilis were isolated and classified into two groups, one of which had resistance to kasugamycin in in vitro protein synthesis and mapped in the ribosomal region . The other group had no resistance to kasugamycin in in vitro protein synthesis and had weak cross-resistance to gentamicin and kanamycin . Neither group could sporulate in the presence of kasugamycin.

J Bacteriol, 1978 Sep, 135(3), 1107 - 17
Genetic and biochemical characterization of kirromycin resistance mutations in Bacillus subtilis; Smith I et al.; Spontaneous mutations causing resistance to the EF-Tu-specific antibiotic kirromycin have been isolated and mapped in Bacillus subtilis . Three-factor transductional and transformational crosses have placed the kir locus proximal to ery-1 and distal to strA (rpsL) and several mutations affecting elongation factors EF-G and EF-Tu, in the order: cysA strA {fus-1/ts-6(EF-G)} {ts-5(EF-Tu)} kir ery-1 spcA . Purified EF-Tu from mutant strains is more resistant to kirromycin as measured by in vitro protein synthesis and also shows a more acidic isoelectric point than wild-type EF-Tu . This indicates that the kir locus is the genetic determinant (tuf) for EF-Tu and that there is a single active gene for this enzyme in B . subtilis.

J Bacteriol, 1978 Sep, 135(3), 1091 - 106
Bacillus subtilis spore coats: complexity and purification of a unique polypeptide component; Goldman RC et al.; Extensively washed, dormant spores of Bacillus subtilis were disrupted with glass beads in buffer at pH 7 in the presence of protease inhibitors . Approximately 31% of the total spore protein was soluble, and another 14% was removed from the insoluble fraction by hydrolysis with lysozyme and washing with 1 M KCl and 0.1% sodium dodecyl sulfate . The residual spore integuments comprised 55% of the total spore proteins and consisted of coats and residual membrane components . Treatment of integuments with sodium dodecyl sulfate and reducing agents at pH 10 solubilized 40% of the total spore protein . Seven low-molecular-weight polypeptide components of this solubilized fraction comprised 27% of the total spore protein . They are not normal membrane components and reassociated to form fibrillar structures resembling spore coat fragments . The residual insoluble material (15% of the total spore protein) was rich in cysteine and was probably also derived from the spore coats . A solubilized coat polypeptide of molecular weight 12,200 has been purified in good yield (4 to 5% of the total spore protein) . Five amino acids account for 92% of its total amino acid residues: glycine, 19%; tyrosine, 31%; proline, 23%; arginine, 13%; and phenylalanine, 6%.

Mikrobiologiia, 1978 Sep-Oct, 47(5), 900 - 5
{Exoprotease properties of Bacillus subtilis var . amyloliquefaciens capable of coagulating blood plasma}; Al'-Nuri MA et al.; The properties of the homogeneous exoprotease preparation from Bacillus subtilis varamyloliquefaciens 759 possessing the coagulase activity were studied . The enzyme is an alkaline protease, has the isopoint at pH 7.8, and not only clots blood plasmo but also hydrolyses such protein substrates as casein, hemoglobin, fibrinogen and fibrin . The enzyme is relatively stable at pH 6.0--9.0 . Bivalent metal ions have virtually no effect on the enzyme activity though some of them stabilize it . The inhibitors PCMB and EDTA do not affect the activity of the enzyme whereas diisopropylfluorophosphate completely inactivates it . Fibrinogen is clotted by the enzyme only in the presence of blood plasma factors.

J Bacteriol, 1978 Sep, 135(3), 1032 - 42
Autolysins and shape change in rodA mutants of Bacillus subtilis; Rogers HJ et al.; The biochemical phenotype of rodA mutants was not affected by the simultaneous presence in double mutants of the lyt gene which makes them 90 to 95% deficient in autolysin action . The only morphological effect of this deficiency on the expression of the rod gene was that both the rod and the coccal forms of the mutant failed to separate and grew as long chains of cells . Inhibition of protein synthesis stopped the increase in peptidoglycan that occurred when the growth temperature for the mutants was raised to 45 degrees C . These observations support the idea that a derepression of peptidoglycan synthesis occurs at this temperature . The increased amount of cellular peptidoglycan at the higher growth temperature is not likely to be the result of the concomitant switching off of autolytic enzyme action.

J Biol Chem, 1978 Aug 25, 253(16), 5585 - 93
Immunochemical studies of the inactivation of aspartate transcarbamylase by stationary phase Bacillus subtilis cells . Evidence for selective, energy-dependent degradation; Maurizi MR et al.; The aspartate transcarbamylase of Bacillus subtilis is stable in exponentially growing cells, but undergoes rapid, energy-dependent inactivation when growth is inhibited by nutrient depletion or addition of antibiotics or other inhibitors of metabolism . This inactivation has been analyzed by a variety of immunochemical techniques, including direct and indirect immunoprecipitation of extracts of cells labeled with 3H-amino-acids, microcomplement fixation, and neutralization of enzymatic activity . The ability of the antibody preparation to react with various denatured, chemically modified, and proteolytically degraded forms of aspartate transcarbamylase was demonstrated . All of the techniques showed that cross-reactive protein disappeared from the cells at the same rate as enzymatic activity, and that little or no immunoprecipitable material of lower than native molecular weight was detectable during inactivation . The disappearance of material cross-reactive with aspartate transcarbamylase occurred prior to the increase in protein degradation that normally occurs in stationary B . subtilis cells and proceeded at a rate at least 20 times greater than general protein degradation . The rate of disappearance was unaffected in mutant strains deficient in intracellular protease activity or in cells treated with inhibitors of protein turnover . Aspartate transcarbamylase was shown to be stable in growing cells . We conclude that the inactivation of aspartate transcarbamylase in vivo involves, or is rapidly followed by, selective, energy-dependent degradation of the protein by a system that appears to involve a previously undescribed protease of B . subtilis.

Biochim Biophys Acta, 1978 Aug 23, 520(1), 82 - 7
DNA synthesis in toluene-treated bacteriophage-infected minicells or Bacillus subtilis; Amann E et al.; Bateriophage (phi29, SPP1, or SPO1)-infected, toluene-treated minicells of Bacillus subtilis are capable of limited amounts of non-replicative DNA synthesis as measured by incorporation of {3H}dTTP into a trichloroacetic acid-precipitable form . The {3H}dTTP is covalently incorporated into small DNA fragments which result from the degradation of a small percentage of the infecting phage genomes (molecular weights in the range of 2 . 10(5)) . Short exposure of the DNA molecules containing the incorporated {3H}dTMP to Escherichia coli exonuclease III results in over 90% of the E13H}dTMP being converted to a trichloroacetic acid-soluble form . The synthesis is totally dependent on host-cell enzymes and is not inhibited by the addition of chloramphenicol, rifampicin, nalidixic acid and mitomycin C and only slightly (approx . 20%) inhibited by the addition of 6-(p-hydroxyphenylazo)-uracil.

Mol Gen Genet, 1978 Aug 17, 164(2), 113 - 29
Mapping by interspecies transformation experiments of several ribosomal protein genes near the replication origin of Bacillus subtilis chromosome; Osawa S et al.; Bacillus subtilis 168 was transformed with DNAs from B . amyloliquefaciens K or B . licheniformis IAM 11054 . These two species show a considerable difference in ribosomal proteins from B . subtilis . Analyses of the transformants indicated that the genes for 16 proteins, S3, S5, S8, S12, S17, S19, BL1, BL5, BL6, BL8, BL14, BL16, BL17, BL22, BL23 and BL25 are located in the cysA-str-spc region on B . subtilis chromosome . The genes for 10 proteins, S4, S6, S13, S16, S20, BL15, BL18, BL20, BL24 and BL28 could not be found in this region in the present experiments.

J Chromatogr, 1978 Aug 11, 155(2), 329 - 36
Phenylboronic acid as a ligand for biospecific chromatography of serine proteinases; Akparov VK et al.; Via attachment of p-(omega-aminoethyl)phenylboronic acid to CH-Sepharose in the presence of water-soluble carbodiimide, a new sorbent for the biospecific chromatography of serine proteinases was obtained . The sorbent was shown to be suitable for the purification of subtilisn, alpha-chymotrypsin and trupsin . It is assumed that the serine hydroxyl group at the active site of the enzyme forms, with the boronic acid moiety of the ligand, a structure that imitates transition enzyme--substrage complex . The presence of glycerol selectively improves the binding of serine proteinases, presumably because of stabilization of the tetrahedral state of the boron atom . Direct isolation of subtilisin from a Bacillus subtilis cultural filtrate on phenylboronic acid-containing sorbent gives a virtually homogenous enzyme (42-fold purification) in a nearly-quantitative yield.

Biochim Biophys Acta, 1978 Aug 7, 525(2), 346 - 56
Two purine nucleoside phosphorylases in Bacillus subtilis . Purification and some properties of the adenosine-specific phosphorylase; Jensen KF; Two purine nucleoside phosphorylases (purine-nucleoside:orthophosphate ribosyltransferase, EC 2.4.2.1) were purified from vegetative Bacillus subtilis cells . One enzyme, inosine-guanosine phosphorylase, showed great similarity to the homologous enzyme of Bacillus cereus . It appeared to be a tetramer of molecular weight 95 000 . The other enzyme, adenosine phosphorylase, was specific for adenosine and deoxyadenosine . The molecular weight of the native enzyme was 153 000 +/- 10% and the molecular weight of the subunits was 25 500 +/- 5% . This indicates a hexameric structure . The adenosine phosphorylase was inactivated by 10(-3) M p-chloromercuribenzoate and protected against this inactivation by phosphate, adenosine and ribose 1-phosphate.

Genetika, 1978 Aug, 14(8), 1303 - 9
{UV-mutagenesis in Bacillus subtilis . VII . Mutability of recA, recB and recF strains}; Filippov VD et al.; UV-light induces reversions to threonine-independence in BD170 Rec+ cells, and does not induce them in isogenic strain BD241 recF . Reversions to methionine-independence are induced in cells GSY1028 recB, and are not in isogenic strain GSY1025 recA . It is possible to construct, with the use of transformation, a viable strain carrying mutations recF and su, the latter imparts threinine-independence to cells . Hence, the absence of UV-induced reversions in recF (and probably in recA) cells can not be explained by the lethal effect of joining, in one genome, two mutations each of which decreases a viability of bacteria . Formation of UV-induced forward mutations which provide additional growth requirements to bacteria is not disturbed in strains recA and recF . Experimental data allow to conclude that UV-induced mutagenesis is not a function of any of two known mechanisms of recombination which function in Bacillus subtilis cells.

Proc Natl Acad Sci U S A, 1978 Aug, 75(8), 3664 - 8
Mechanism of integrating foreign DNA during transformation of Bacillus subtilis; Duncan CH et al.; Genes encoding thymidylate synthetase from Bacillus subtilis bacteriophages were cloned in Escherichia coli . Chimeric plasmids pCD1 and pCD3 were constructed from site-specific endonuclease digests of bacteriophage phi3T DNA cloned in pMB9 in E . coli . Similar cloning techniques with bacteriophage beta22 DNA yielded chimeric plasmids pCD4, pCD5, and pCD6 . Endonuclease digests of DNA from pCD1 and pCD3 propagated in E . coli or from DNA isolated from bacteriophage phi3T propagated in B . subtilis transformed B . subtilis from Thy- to Thy+ . Intact DNA from bacteriophage beta22, endonuclease digests of beta22 DNA, and a chimeric plasmid (pCD5) composed only of the thybeta22 gene and pMB9 did not transform B . subtilis from Thy- to Thy+ even though pCD5 could transform Thy- E . coli to Thy+ . However, if the thybeta22 fragment from pCD5 was introduced into another chimeric plasmid, pCD2, that contains a region of homology to the chromosome of B . subtilis in addition to pMB9, transformation of Thy- clones of B . subtilis was possible . Furthermore, Southern hybridization analyses of the digests of chromosomal DNA from the Thy+ transformants established that the entire chimeric plasmid was incorporated into the chromosome of B . subtilis . Treatment of these plasmids with site-specific endonucleases abolished transformation . These results indicated that the entire chimeric plasmid can be incorporated into the chromosome of B . subtilis by a Campbell-like model . Therefore, an additional mechanism for transformation exists whereby plasmids can be integrated if sufficient chromosomal homology is maintained.

J Bacteriol, 1978 Aug, 135(2), 408 - 14
Pleiotropic effect of a rifampin-resistant mutation in Bacillus subtilis; Ryu JI; Rifampin-resistant (Rifr) mutants were isolated spontaneously from Bacillus subtilis strain 168 . A fraction of the mutants did not grow on a minimal medium . A high concentration of one of the L-amino acids (glutamic acid, glutamine, arginine, proline, aspartic acid, or asparagine) was required to restore their growth on the medium . Further analysis of one of the mutants (strain RF 161) suggested that the mutant is unable to use ammonia as a nitrogen source and requires amino acids instead . Activity of glutamate synthase was not detected in the crude extract of the mutant . The Rifr mutation was closely located to cysA and the drug resistance was cotransformed with the property of amino acid requirement at 100% frequency . All revertants to prototrophy tested showed the rifampin-sensitive (Rifs) property . The activity of the DNA-dependent RNA polymerase of the mutant was resistant to rifampin . It is concluded that some alteration of RNA polymerase may cause absence of the activity of an enzyme involved in the nitrogen metabolism.

J Biol Chem, 1978 Jul 25, 253(14), 4999 - 5004
Dehydroquinate synthase in Bacillus subtilis . An enzyme associated with chorismate synthase and flavin reductase; Hasan N et al.; Dehydroquinate synthase, the enzyme which catalyzes the conversion of 3-deoxy-D-arabino-heptulosonic acid 7-phosphate (DAHP) to 5-dehydroquinate, has been purified from Bacillus subtilis in association with chorismate synthase and NADPH-dependent flavin reductase . The enzyme was only active when associated with chorismate synthase, whereas the flavin reductase could be separated from the complex with retention of dehydroquinate synthase activity . The enzyme requires NAD and either Co2+ or Mn2+ for maximal activity . The activity was completely inhibited by EDTA . The Km of the enzyme for DAHP, NAD, and Co2+ were estimated to be 1.3 X 10(-4), 5.5 X 10(-5), and 5.5 X 10(-5) M, respectively . Enzyme activity was completely inhibited by NADH and the inhibition was not reversed by the addition of NAD, NADPH and NADP were not inhibitory . The enzyme was unstable to heat and lost all activity at 55 degrees C . A protein fraction which did not adsorb to phosphocellulose was found to inhibit the enzyme.

J Biol Chem, 1978 Jul 25, 253(14), 4993 - 8
Purification and properties of chorismate synthase from Bacillus subtilis; Hasan N et al.; Chorismatic synthase was purified to apparent homogeneity from Bacillus subtilis . The enzyme required NADPH-dependent flavin reductase, Mg2+, NADPH, and flavin (FMN or FAD) for activity . The molecular weight of chorismate synthase was 24,000 as determined by sodium dedecyl sulfate (SDS)-gel electrophoresis . The enzyme was also isolated in a complex form associated with NADPH-dependent flavin reductase and another enzyme of the aromatic amino acid pathway, dehydroquinate synthase . On SDS-gel electrophoresis, this form was resolved into three bands with molecular weights of 13,000, 17,000, and 24,000 . The enzyme complex was easily dissociated and the dissociation resulted in a change in the chromatographic properties of NADPH-dependent flavin reductase which was no longer retained on phosphocellulose whereas chorismate synthase was still adsorbed . Chorismate synthase activity was linear with time and protein concentration, whereas partially purified preparations showed a significant lag period before the reaction took place . Moreover, crude or partially purified enzyme preparations were completely inactivated by dilution and the activity could be recovered by addition of flavin reductase . A possible role of NADPH-dependent flavin reductase in the activation and regulation of chorismate synthase activity is discussed.

J Biol Chem, 1978 Jul 25, 253(14), 4987 - 92
Purification and characterization of NADPH-dependent flavin reductase . An enzyme required for the activation of chorismate synthase in Bacillus subtilis; Hasan N et al.; NADPH-dependent flavin reductase (required for the activation of chorismate synthase) was purified to homogeneity from cell-free extracts of Bacillus subtilis . The enzyme has a molecular weight of 13,000 as determined by sodium dodecyl sulfate-gel electrophoresis, is specific for NADPH, and requires a divalent metal ion and either FMN or FAD for maximal rates of NADPH oxidation . The enzyme is able to reduce 2,6-dichlorophenolindophenol (DCIP) in the presence of NADPH and a divalent metal ion . Both catalytic activities were completely inhibited by EDTA . The Km for FMN is 1.25 X 10(-5) M and for NADPH 7.8 X 10(-5) M with oxygen as the final electron acceptor, and 3.85 X 10(-4) M with DCIP as the final electron acceptor . The enzyme was also isolated in association with chorismate synthase and dehydroquinate synthase . The enzyme associated with the complex has the same catalytic properties as the dissociated enzyme except that it requires both a divalent metal ion and FMN for DCIP reduction . Maximal enzyme activity was observed when the enzyme was preincubated with FMN and the divalent metal ion . The enzyme complex is easily dissociable and the dissociation of the enzyme complex resulted in the failure of NADPH-dependent flavin reductase to adsorb to phosphocellulose.

J Biol Chem, 1978 Jul 25, 253(14), 4916 - 9
Independence of proline chemotaxis and transport in Bacillus subtilis; Ordal GW et al.; Values of KI for nine proline analogs as inhibitors of proline chemotaxis and of proline transport were determined . Two of them inhibited transport at substantially lower concentrations than chemotaxis; two at substantially higher concentrations . Moreover, mutants, believed to be in the component that binds proline, were isolated that showed a shift of KM for transport to higher concentrations, one as much as 40-fold . However, chemotaxis was virtually unaffected . Therefore, unlike galactose chemotaxis and transport in Escherichia coli, which share the galactose-binding protein, proline chemotaxis and transport in Bacillus subtilis are independent.

Biochim Biophys Acta, 1978 Jul 24, 519(2), 513 - 25
Studies on the negative circular dichroic bands around 297 nm of ribosomes from bacterial cells; Uchiumi T et al.; The small negative CD bands around 297 nm of isolated 30-S and 50-S ribosomal subunits were precisely measured for three bacteria, Bacillus stearothermophilus, Bacillus subtilis and Escherichia coli Q 13 . The intensities of the negative CD bands of 30-S subunits were always much greater than those of 50-S subunits irrespective of the bacterial strains, which may be related to the difference in comformations of rRNAs and proteins in the complexes between these subribosomal particles . The dissociation of 70-S ribosomes into two subunits by lowering Mg2+ concentration caused evident enhancement of intensity of the 297 nm CD band, which was completely reversed by the association of the two subunits into 70-S particles . The melting profiles of CD spectra 3 B . stearothermophilus and E . coli were compared and both subunits of the former were found to be more heat stable than those of the latter . It was found that 5 M urea and 0.5% sodium dodecyl sulfate (SDS) treatment caused considerable reduction of the negative CD intensity around 297 nm of 30-S subunits but no significant change of 50-S subunits, while no significant change was observed for the CD spectra of isolated 16-S and 23-S rRNAs by the same treatment . Effects of EDTA treatment and then addition of Mg2+ on the CD spectra and fluorescence emission spectra of the subunits were also observed and the contribution by the interaction between rRNA s and proteins in ribosomes to the small negative band around 297 nm was discussed.

Mol Gen Genet, 1978 Jul 6, 163(1), 7 - 15
Transfection with replicating DNA from the temperate Bacillus bacteriophage phi 105 and with T4-ligase treated phi105 DNA: the importance in transfection of being longer than genome-length; Flock JI; Replicating phage DNA extracted from Bacillus subtilis infected with phage phi 105 has a higher activity in transfection than mature DNA . By heteroduplex analysis it was shown that this DNA contains concatemeric molecules . Concatemers, constructed in vitro by treatment of mature DNA with T4-ligase also have an increased activity in transfection . DNA showing an increased activity in transfection does not have a requirement for more than one molecule per transfection event as is typically found for transfection with mature phi 105 DNA . An explanation is given for this difference suggesting that the structure of the ends of the transfecting molecules play an important role intransfection.

J Virol, 1978 Jul, 27(1), 255 - 7
Dextran sulfates as a contaminant of DNA extracted from concentrated viruses and as an inhibitor of DNA polymerases; Hitzeman RA et al.; Dextran sulfate is commonly used with polyethylene glycol to concentrate viruses before extraction of their DNA . However, dextran slulfate then easily contaminated such DNA and acted as a potent inhibitor of DNA polymerases from Bacillus subtilis (III), phage PBS2, and phage T4 . Dextran sulfate only weakly inhibited Micrococcus luteus and Escherichia coli DNA polymerase I preparations.

Mikrobiologiia, 1978 Jul-Aug, 47(4), 717 - 21
{Electron microscopic study of Bacillus subtilis mutants differing in the serine proteinase activity and spectrum}; Smirnov TA et al.; Vegetative cells and spores of the colonial morphological mutants of Bacillus subtilis A-50 were studied by electron microscopy . The ultrastructure of vegetative cells from both asporogenic colonial-morphological mutants and those which were capable of forming spores in the presence of high concentrations of nitrogen and carbon sources with a decreased activity and a modified spectrum of serine proteases differed from the parent strain by the presence of a microcapsule, the uneven thickness of a cell wall, and the absence of a distinct periplasmic space . Crystalline inclusions of a regular shape were detected in the sporeforming mutant in those cells which were devoid of spores . Spores of the mutant had additional layers.

Arch Microbiol, 1978 Jul, 118(1), 27 - 34
Uracil incorporation in the forespore and the mother cell during spore development in Bacillus subtilis . Autoradiographic electron microscopic study; Ryter A et al.; The transcriptional activity of the two genomes of the sporangium during spore formation was determined by pulse-labeling bacteria with 3H-uracil at different times of sporulation and preparing them for high resolution autoradiography . The quantitative analysis of autoradiographs shows that uracile incorporation in the whole sporangium decreases considerably between stages II and IV . However, the variations of the transcriptional activity are not identical in the mother cell and in the forespore . The one of the mother cell decreases rapidly between stages II and III and then remains stable until the end of stage IV, whereas that of the forespore which is low at stage II increases as the forespore grows ovoid and then quickly diminishes . It is very weak at the beginning of stage IV and negligible at the end of this stage . Pulse-chase experiments made in the presence of rifampine indicate that about 30% of the uracile incorporated is located in stable RNA . This value is found at any stage of sporulation in both cellular compartments whatever their rate of uracile incorporation . A relationship can be made between the nuclear shape and the activity of the genetic material . This confirms observations made by several authors in other bacterial species and other physiological conditions that the condensed shape corresponds to a state of low transcriptional activity whereas the more irregular and dispersed shape corresponds to a state of high activity.

J Microsc, 1978 Jul, 113(2), 131 - 8
Signal to noise enhancement in a study of cell wall structure of Bacillus subtilis by interactive computation; Barrett AN et al.; This paper describes a technique for obtaining values of the width and variation in optical density of sectioned bacterial cell walls by interactive computational methods . Background 'noise' prevents accurate determination of cell wall boundaries from the data in a single scan line but the noise may be suppressed by averaging several consecutive scan lines . although application of the technique is explained for sectioned bacterial cell walls, it is equally valid for similar situations where single line scans of electron micrographs are inadequate for precise determination of measurement.

J Med Chem, 1978 Jul, 21(7), 607 - 12
Synthesis and evaluation of bis-dipeptide and bis-tripeptide analogues of actinomycin D; Azad Chowdhury AK et al.; Six-bis-dipeptide analogues of actinomycin D, all containing two threonyl-D-valine side chains, were prepared . Also two bis-tripeptide analogues containing an additional proline or oxoproline residue were synthesized . None of the compounds bound to DNA in a manner similar to actinomycin D . This lack of strong intercalative binding emphasizes the importance of the pentapeptidolactone side chains in the binding of actinomycin D to DNA and also highlights the deficiences inherent in using only small nucleotide sequences in investigating drug-DNA binding . None of the analogues tested showed any antitumor activity, although actinocylbis(threonyl-D-valine methyl ester) did show 10% of the antibacterial activity of actinomycin D vs . Bacillus subtilis.

J Bacteriol, 1978 Jul, 135(1), 99 - 113
Evidence that spo0A mutations are recessive in spo0A-/spo0A+ merodiploid strains of Bacillus subtilis; Trowsdale J et al.; The spo0A locus contains two types of closely linked mutations that block sporulation at stage 0: spo0A mutations (the most pleiotropic of the stage 0 markers) and spo0C mutations . It was previously thought that spo0A mutations were dominant in merodiploids of Bacillus subtilis, whereas spo0C mutations were recessive . We have shown that spo0A mutations were recessive when spo0A-/spo0A+ merodiploids were made in the genetic backgrounds of strains that were resistant to antibiotic produced by the wild-type strain . Reinvestigation of cultures of spo0A-/spo0A+ merodiploids constructed in the antibiotic-sensitive spo0A strain showed that they contained the spo0A allele at a low frequency, and they produced very few haploid Spo- segregants . These facts indicated that the cultures contained mostly homogenotic (spo0A+/spo0A+) cells . The reason for the poor survival of the spo0A-/spo0A+ merodiploids in the genetic background of the antibiotic-sensitive strain was not clear, but several possible explanations were given . It may have been related to the diploid state of other genes in the same merodiploid cells . The previous indication that spo0A mutations were dominant seems to have been based on the properties of a rare class of Spo- segregants that were probably selected in the presence of antibiotic.

Science, 1978 Jun 30, 200(4349), 1493 - 4
Thermorestoration of mutagenic radiation damage in bacterial spores; Tanooka H; The frequency of mutations in Bacillus subtilis spores irradiated anoxically with ionizing radiation decreased as a result of subsequent anoxic heating to the same extent as the increase in spore survival . This result indicates that DNA damage is the origin of the "thermorestoration" phenomenon.

Biochim Biophys Acta, 1978 Jun 8, 502(3), 445 - 57
Membrane ATPase of Bacillus subtilis . I . Purification and properties; Serrahima-Zieger M et al.; The membrane ATPase (EC 3.6.1.3) of Bacillus subtilis can be solubilized by a shock-wash process . Two procedures for purifying the solubilized enzyme are reported . A protease inhibitor, phenylmethane sulfonylfluoride, was introduced in the solubilization and purification step . The resultant ATPase purified by density gradient centrifugation has a molecular weight of 315 000, an s20,w of 13,4 and an amino acid composition very similar to bacterial ATPases already studied . After exposure to polyacrylamide gel electrophoresis in presence of sodium dodecyl sulphate (SDS), or 8 M urea or SDS-urea, the purified ATPase can be dissociated in two non-identical subunits of molecular weights 59 000 (alpha) and 57 000 (beta) with different charges . Kinetic studies showed that Ca2+ or Zn2+ are required for ATPase activity, although Mg2+ was uneffective . At optimal Ca2+ concentration, the Mg2+ has an inhibitory effect . The Km for ATP is 1.3 mM . Inhibitors of the oxydative phosphorylation, of the mitochondrial ATPase and of the (Na+ + K+)-ATPase are studied.

J Bacteriol, 1978 Jun, 134(3), 920 - 8
Biochemical and genetic study of D-glucitol transport and catabolism in Bacillus subtilis; Chalumeau H et al.; The catabolic pathway of D-glucitol (sorbitol) in Bacillus subtilis Marburg 168M is characterized . It includes (i) a transport step catalyzed by a D-glucitol permease which is affected by the gutA mutations, (ii) an oxidation step of the intracellular D-glucitol catalyzed by a D-glucitol dehydrogenase, generating intracellular fructose, affected by gutB mutations, and (iii) phosphorylation of the intracellular fructose either at the C1 site or at the C6 site as described previously (A . Delobbe et al., Eur . J . Biochem., 66:485-491, 1976; A . Delobbe et al., EUR . J . Biochem . 51:503-510, 1975) . Additional data are given concerning the phosphorylation of fructose by a fructokinase (fructose ATP 6-phosphotransferase), which is affected by the fruC mutation . The isolation of regulatory mutants affected in gutR that synthesize constitutively both the permease and the dehydrogenase indicates the existence of a D-glucitol operon in B . subtilis . Unlike the wild-type strain, these mutants are able to utilize D-xylitol as sole carbon source.

Infect Immun, 1978 Jun, 20(3), 782 - 91
Lysozyme: primary bactericidin in human plasma serum active against Bacillus subtilis; Selsted ME et al.; The in vitro bactericidal reaction of human plasma serum against Bacillus subtilis was investigated . Human lysozyme was purified to homogeneity, and antiserum was prepared against the enzyme . The anti-lysozyme immunoglobulin G was used as a specific inhibitor in bactericidal and bacteriolytic reactions . It was found that at low serum concentrations lysozyme was the primary bactericide active against B . subtilis . At appreciably higher serum concentrations, a lysozyme-independent bactericidal activity was also demonstrated.

J Bacteriol, 1978 Jun, 134(3), 973 - 81
Bacillus subtilis 168 genetic transformation mediated by outgrowing spores: necessity for cell contact; Orrego C et al.; Transforming activity released in sequential genetic order during the first synchronous cycle of DNA replication during outgrowth of spores of Bacillus subtilis 168 was investigated . A transformation assay was used consisting of outgrowing spores as DNA donors and multiply marked competent cells as recipients . DNA synthesis inhibitors known to stop DNA release were used during and subsequent to DNA transfer to recipient cells . The released DNA sedimented with the outgrowing cells after low-speed centrifugation, and it was discovered that markers released both early and late were resistant to up to 500 microgram of deoxyribonuclease per ml under conditions in which the transforming capacity of purified DNA was eliminated by 5 microgram of the nuclease per ml . Inaccessibility to deoxyribonuclease was increased and maintained during the transformation event while detergents and proteolytic attack did not expose the released chromosome to nuclease action . The results indicate that tight physical contact between outgrowing spores and competent cells is required for transformation in this system.

J Biol Chem, 1978 May 25, 253(10), 3536 - 42
Deoxycytidylate deaminase from Bacillus subtilis . Purification, characterization, and physiological function; Mollgaard H et al.; dCMP deaminase from Bacillus subtilis has been purified 700-fold . In addition to the substrate, dCMP, the enzyme requires dCTP, Zn2+, and 2-mercaptoethanol, Mg2+ cannot substitute for Zn2+ . The dCMP saturation curve is hyperbolic in the presence of saturating concentrations of dCTP and Zn2+ . The dCTP saturation curve is sigmoidal, the sigmoidicity being dependent on the Zn2+ and dCMP concentrations . The molecular weight as determined by gel filtration is 170,000 both in the presence and in the absence of dCTP and Zn2+ . In the absence of thiols, the enzyme is highly unstable . At 0 degrees, the half-life of the enzyme activity is 30 min . Addition of Zn2+ and dCTP protects against this inactivation . In the presence of a thiol, dCTP and Zn2+ protect the enzyme against heat inactivation at 50 degrees . A mutant lacking dCMP deaminase (dcd) was isolated . Labeling of the pyrimidine nucleotide pools reveals that in the parent strain, 45% of the dTTP pool is derived via dCMP deamination, the residual 55% being derived via reduction of a uridine nucleotide . Since the dcd mutant grows with the same doubling time as the parent strain, we conclude that uridine nucleotide reduction alone is capable of supplying sufficient dUMP for normalthymidine nucleotide synthesis.

Eur J Biochem, 1978 May 16, 86(2), 531 - 7
Use of the T4 polynucleotide ligase in the joining of flush-ended DNA segments generated by restriction endonucleases; Sgaramella V et al.; Double-stranded DNA segments with completely base-paired ends were obtained by the action of various restriction endonucleases on phage and plasmid DNAs . These segments were joined covalently by the T4 polynucleotide ligase . The joining was monitored by the electron microscopy count of intramolecularly circularized segments . The highest extent of joining, close to 75%, was observed at 15-25 degrees C with the segments resulting from the action of the Bacillus subtilis (strain R) restriction endonuclease Bsu on the DNA of bacteriophage SPPI or of the plasmid pSC 101 . The joining of double-stranded termini required about 10 times more enzyme than the short single-stranded termini produced by the Escherichia coli restriction endonuclease EcoRI . A shortened purification of the T4 ligase was found to give an enzyme devoid of interfacing nucleases.

Biochim Biophys Acta, 1978 May 11, 524(1), 60 - 7
Aminotransferases for aromatic amino acids and aspartate in Bacillus subtilis; Mavrides C et al.; Two proteins (form A and form B2) with aromatic-amino-acid aminotransferase activity were detected in extracts of Bacillus subtilis . A histidinol phosphate aminotransferase (protein B1) with aminotransferase activity for the aromatic amino acids was also present . The aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1) (protein C) also displayed similar activity . Each of the four proteins was isolated free from the others by the successive application of DEAE-cellulose column chromatography and flat-bed isoelectric focusing at pH range 4-6 . Form B2 is the major form of the aromatic-amino-acid aminotransferase (aromatic-amino-acid:2-oxoglutarate amino-transferase, EC 2.6.1.57) and the Km values of tyrosine and phenylalanine with this form are somewhat lower than with the minor form A . The Km of tyrosine with histidinol phosphate aminotransferase (protein B1) is in the same range, but the Km of phenylalanine with this enzyme is 12-20 times higher than the corresponding values with the two forms of the aromatic-amino-acid amino-transferase . Apparent molecular weights were estimated with Sephadex gel filtration to be approx . 73 000, 64 000, 54 000 and 66 000 for form A, form B2, histidinol phosphate aminotransferase and aspartate aminotransferase, respectively . Form B2 is being reported for the first time in this communication.

Mol Gen Genet, 1978 May 3, 161(2), 135 - 41
Delta factor can displace sigma factor from Bacillus subtilis RNA polymerase holoenzyme and regulate its initiation activity; Williamson VM et al.; A protein with a molecular weight of 21,000 daltons is found associated with a fraction of Bacillus subtilis RNA polymerase core . This protein (delta) does not react with antibody made against sigma factor and has a peptide map which is significantly different from sigma factor . At ratios of 2:1 to 4:1 (delta:holoenzyme) the delta displaces sigma factor completely from the core and associates in a 1:1 ratio with core to form delta-core . Under the same incubation conditions sigma factor at a ratio of 10:1 (sigma factor:delta-core) does not displace delta from the delta-core . The delta-core has much less activity as compared to holoenzyme on various DNA templates . However, sigma factor does stimulate the activity of delta-core enzyme under conditions of RNA synthesis . These observations and the results of others suggest that delta-core enzyme binds initially to specific DNA sites followed by delta release from the core-DNA complex and that the sigma factor binds to the core-DNA complex to initiate RNA synthesis . Thus both delta and sigma factors are required in a sequential fashion for specific transcription to occur in B subtilis.

Biochemistry, 1978 May 2, 17(9), 1785 - 91
Identification of phyllostine as an intermediate of the patulin pathway in Penicillium urticae; Sekiguchi J et al.; A patulin negative mutant (J1) of Penicillium urticae (NRRL 2159A) was found to accumulate large quantities (greater than 128 mg/L culture) of a reactive, photosensitive compound, which was isolated and identified as (-)-phyllostine (5,6-epoxygentisylquinone) . This epoxyquinone possessed an antibiotic activity against Bacillus subtilis which was approximately 80% of that exhibited by patulin . In separate in vivo feeding experiments, {2-14C}acetate and {G-3H}gentisaldehyde were readily incorporated into phyllostine by mutant J1 and {14C}phyllostine was incorporated into patulin by the parent strain (NRRL 2159A) . When fed to a washed-cell suspension of a second patulin negative mutant (J2) which produced gentisaldehyde but not phyllostine, unlabeled phyllostine was efficiently converted to patulin in yields of 33, 56, and 92% after 30 min, 1 and 5 h, respectively . The role of phyllostine as an intermediate of a new post-gentisaldehyde portion of the patulin biosynthetic pathway is discussed.

J Bacteriol, 1978 May, 134(2), 680 - 2
Outgrowth and vegetative growth inhibition of Bacillus subtilis spores after transient exposure to polymyxin B; Tochikubo K; Polymyxin B combined with the resting spores of Bacillus subtilis and inhibited outgrowth and vegetative growth after germination . The antibiotic was released from the resting spores and its inhibitory action was reversed by the addition of di- and trivalent metallic cations.

Ann Microbiol (Paris), 1978 May-Jun, 129(4), 525 - 43
{Induction of interferon by Bacillus subtilis (author's transl)}; Nozaki-Renard J; Although it is generally admitted that Gram-positive bacteria cannot induce interferon, we were able to show that organisms and cell walls of Bacillis subtilis as well as components obtained through the extraction of endotoxins from Gram-negative bacteria can induce type II interferon . Moreover we have ascertained that those components have the same biological activity as the endotoxins, espcially with regard to their behaviour with B cells of lymphocytes, the results of pyrogen test, the incidence of cycloheximide and their function as inhibitors of bacteriophages . However, it has appeared that chemically they are quite different from the lipopolysaccharide of the Gram-negative bacteria: no lipid A could be detected.

Hoppe Seylers Z Physiol Chem, 1978 May, 359(5), 559 - 70
{Vitamin B 6 biosynthesis in Bacillus subtilis}; Pflug W et al.; 1) A vitamin-B6-producing mutant, BA 1, was selected by treatment of Bacillus subtilis with N-methyl-N'-nitro-N-nitrosoguanidine . Using gradient plates supplemented with the vitamin B6 antagonist isonicotinohydrazide, three mutants of BA 1 were isolated, which excrete 2-5 mg of vitamin B6/l of growth medium . 2) Mutation of the three vitamin-B6-producing strains BA 1, BA 11 and L 71 led to the isolation of 49 vitamin-B6 deficient mutants . All mutants are able to grow with pyridoxine, pyridoxal, pyridoxamine, and even with 4'-deoxypyridoxine . Glycolaldehyde or nicotinic acid do not support growth of the mutants . Some of these vitamin-B6-deficient mutants can also grow in the absence of vitamin B6, providing isoleucine is present . Others show a growth stimulation, when isoleucine is added to a medium containing a vitamin B6 compound . Isoleucine can be replaced by 3-methyl-2-oxovalerate . Cross-feeding experiments indicated a division of the mutants into two groups . Using chromatographic methods, substances which support growth of the mutants were purified, but have not yet been identified . Following the addition of 4'-deoxypyridoxine, 4'-deoxypyridoxine 5'-phosphate was isolated from the growth medium of a vitamin B6-deficient mutant . 3) Threonine dehydratase, transaminase B and transaminase C from wild-type Bacillus subtilis were compared with the enzymes from vitamin-B6-producing strains and with the enzymes from vitamin-B6-deficient mutants . Both the vitamin-B6-producing and the vitamin B6-deficient mutants show higher specific activities than wild type . In the mutant strains no multivalent repression of the threonine dehydratase and transminase B by isoleucine, leucine and valine could be demonstrated . Leucine dehydrogenase, the first enzyme of the isoleucine catabolic pathway, is constitutively produced in the vitamin-B6-producing and in the vitamin-B6-deficient mutants . In the vitamin-B6-deficient mutants, there is a correlation between growth yield in the presence of isoleucine and the specific activity of leucine dehydrogenase . In the crude extract of Bacillus subtilis no pyridoxamine-phosphate oxidase activity could be demonstrated, whereas pyridoxal kinase was readily detectable.

J Biochem (Tokyo), 1978 May, 83(5), 1503 - 10
Tryptophan residues of saccharifying alpha-amylase from Bacillus subtilis . A kinetic discrimination of states of tryptophan residues using N-bromosuccinimide; Fujimori H et al.; Four tryptophan residues of saccharifying alpha-amylase from B . subtilis out of eleven in total are reactive towards N-bromosuccinimide (NBS), suggesting that they are on the surface of the enzyme . This is consistent with the results of solvent perturbation difference spectrophotometry with ethylene glycol . One of four tryptophan residues was clearly distinguished from the other three in reactivity with NBS by the stopped-flow method . This most reactive tryptophan residue was not protected from modification by substrates of analogs, indicating that the tryptophan is not located in the substrate binding site . One of the other three tryptophan residues, probably the second most reactive one, is considered to be related in some way to the glycosyl transfer in the reaction of the enzyme with maltose as a substrate.

J Bacteriol, 1978 May, 134(2), 514 - 20
Recombination between compatible plasmids containing homologous segments requires the Bacillus subtilis recE gene product; Keggins KM et al.; Plasmid pSL103 was previously constructed by cloning a Trp fragment (approximately 2.3 X 10(6) daltons) from restriction endonuclease EcoRI-digested chromosome DNA of Bacillus pumilus using the neomycin-resistance plasmid pUB110 (approximately 2.8 X 10(6) daltons) as vector and B . subtilis as transformation recipient . In the present study the EcoRI Trp fragment from pSL103 was transferred in vitro to EcoRI fragments of the Bacillus plasmid pPL576 to determine the ability of the plasmid fragments to replicate in B . subtilis . Endonuclease EcoRI digestion of pPL576 (approximately 28 X 10(6) daltons) generated three fragments having molecular weights of about 13 X 13(6) (the A fragment), 9.5 X 10(6) (B fragment, and 6.5 X 10(6) (C fragment) . Trp derivatives of pPL576 fragments capable of autonomous replication in B . subtilis contained the B fragment (e.g., pSL107) or both the B and C fragments (e.g., pSL108) . Accordingly, the B fragment of pPL576 contains information essential for autonomous replication . pSL107 and pSL108 are compatible with pUB110 . Constructed derivatives of the compatible plasmids pPL576 and pUB110, harboring genetically distinguishable EcoRI-generated Trp fragments cloned from the DNA of a B . pumilus strain, exhibited relatively high frequency recombination for a trpC marker when the plasmid pairs were present in a recombination-proficient strain of B . subtilis . No recombination was detected when the host carried the chromosome mutation recE4 . Therefore, the recE4 mutation suppresses recombination between compatible plasmids that contain homologous segments.

J Bacteriol, 1978 May, 134(2), 476 - 82
Biosynthesis of riboflavin in Bacillus subtilis: function and genetic control of the riboflavin synthase complex; Bacher A et al.; Two riboflavin synthase activities (heavy and light) have been observed in earlier studies with Bacillus subtilis . The heavy enzyme is a complex of one molecule of light enzyme (consisting of three alpha subunits) and approximately 60 beta subunits (A . Bacher, R . Bauer, U . Eggers, H . Harders, and H . Schnepple, p . 729--732, in T . P . Singer (ed.), Flavins and Flavoproteins, Elsevier, Amsterdam, 1976) . The formation of alpha and beta subunits is coordinately controlled . Mutants apparently deficient in beta subunits were isolated as riboflavin requires after mutagenesis of B . subtilis with ICR 191 . The mutants could grow with diacetyl instead of riboflavin . Growth with diacetyl was associated with the accumulation of substantial amounts of the riboflavin precursor, 6,7-dimethyl-8-(D-ribityl)lumazine . It follows that the mutants are deficient in an enzyme activity required for the formation of the lumazine from the pyrimidine precursor . We conclude that heavy riboflavin synthase is a bifunctional enzyme . The riboflavin synthase activity is mediated by the alpha subunits, whereas the beta subunits are necessary for an earlier biosynthetic step.

J Bacteriol, 1978 May, 134(2), 440 - 5
Early-blocked asporogenous mutants of Bacillus subtilis are lysogenized at reduced frequency by temperate bacteriophages; Ikeuchi T et al.; The establishment of lysogeny in early-blocked asporogenous (Spo-) mutants of Bacillus subtilis 168, which were also defective in the production of antibiotics (Abs-), by temperate phage phi105 or SPO2 was studied . It was found that the frequency of lysogenization of Spo-Abs-mutants was 10 to 20% that of the wild-type bacteria . There was no difference in the efficiency of plating and the burst size of phi105 between wild-type and mutant strains . Phi105 lysogens of mutant strains were as stable as those of the wild type . Several rifampin-resistant mutants defective in the production of antibiotics were isolated . They were also defective in spore formation and lysogenized by phi105 at reduced frequency.

Can J Microbiol, 1978 May, 24(5), 563 - 8
Induction of autolysis in Bacillus subtilis by ochratoxin A; Singer U et al.; Ochratoxin A (OTA) added during the exponential growth phase at a concentration higher than 12 microgram/ml caused autolysis of Bacillus subtilis . Optical density of cultures decreased, and at higher concentrations the cultures became sterile . Optimum OTA-induced lysis was about pH 5 . At concentrations below 10 microgram/ml, protein synthesis was inhibited more strongly than RNA synthesis . Cell wall synthesis was also strongly inhibited . A fraction extracted from the lysates had the property of a lysis inhibitor . The relevance of this fraction in respect to autolysis is discussed.

Biochim Biophys Acta, 1978 Apr 19, 540(1), 48 - 54
Biosynthesis of riboflavin . 6,7-Dimethyl-8-ribityllumazine 5'-phosphate is not a substrate for riboflavin synthase; Harzer G et al.; Phosphotransferase from carrot is shown to catalyze the phosphorylation of 6,7-dimethyl-8-ribityllumazine specifically at position 5' of the ribityl side chain . The lumazine 5'-phosphate is neither a substrate nor an inhibitor of riboflavin synthase from Bacillus subtilis and Escherichia coli . It follows that the obligatory product of riboflavin synthase is riboflavin and not FMN.

Eur J Biochem, 1978 Apr 17, 85(2), 433 - 6
Further evidence for the structure of the teichoic acids from Bacillus stearothermophilus B65 and Bacillus subtilis var . niger WM; de Boer WR et al.; Bacillus stearothermophilus B65 and Bacillus subtilis var . niger WM both contain teichoic acids in their walls composed of glycerol, phosphate and glucose . The 13C nuclear magnetic resonance spectrum of B . stearothermophilus teichoic acid showed 13C-31P coupling on the signals from the C-5 and C-6 carbon atoms of the glucose molecule and an alpha-glucosidic linkage between glucose and the C-1 atom of the glycerol moiety . These data are consistent with a poly{glucosylglycerol phosphate} as the cell-wall teichoic acid in this organism . B . subtilis var . niger WM teichoic acid was oxidized by periodate and incubated in glycine buffer at pH 10.5 . This treatment did not significantly increase the phosphomonoester content (by beta-elimination of the phosphate groups) of the teichoic acid molecule (7.1 to 9.5%), which is in accordance with earlier data derived from 13C nuclear magnetic resonance spectroscopy {De Boer et al . (1976) Eur . J . Biochem . 62, 1-6}, that in this organism the glucose is not an integral part of the polymer chain . Similar treatment of B . stearothermophilus B65 teichoic acid increased the phosphomonoester content of the preparation from 0.15 to 68.1%.

Biochem J, 1978 Apr 15, 172(1), 63 - 7
Extracellular manganese-stimulated deoxyribonuclease as a marker event in sporulation of Bacillus subtilis; Akrigg A et al.; A considerable amount of Mn2+-stimulated DNAase (deoxyribonuclease) activity is released by Bacillus subtilis 168 during sporulation in a glucose-deficient medium; much smaller amounts are released during starvation for phosphate or nitrogen . Protein synthesis is required . Two forms of evidence are presented that production of the DNAase is associated with events late in stage II of sporulation . 19 Thymidine starvation, which inhibits the biochemical events associated with sporulation, also inhibits release of the DNAase . 2 . Several asporogenous mutants blocked at stage II or earlier and unable to produce alkaline phosphatase (a stage-II event) do not produce the enzyme . Mutants blocked towards the end of stage II or later produce both enzymes . During sporulation of the wild-type strain, the DNAase appears about 1 h after alkaline phosphatase . The results suggest that production of the DNAase is controlled by a still-undiscovered stage-II genetic locus.

Biochem J, 1978 Apr 15, 172(1), 69 - 76
Purification and properties of a manganese-stimulated deoxyribonuclease produced during sporulation of Bacillus subtilis; Akrigg A; A DNAase (deoxyribonuclease) was isolated from culture supernatants of sporulating Bacillus subtilis 168 . The purified enzyme migrated as a single band during polyacrylamide-gel electrophoresis . The enzyme differs from other DNAases of B . subtilis in molecular weight, metal-ion requirement and mode of action . The enzyme was inactive in the absence of metal ions, and exhibited optimum activity with 10 mM-Mn2+, although Mg2+, Cd2+ and Co2+ could also permit some activity . The pH optimum for the enzyme was pH 7.5, and it degraded linear-duplex DNA or closed-circular-duplex DNA to acid-soluble material . There was little or no activity on single-stranded DNA or rRNA . Sucrose-gradient analysis of the products of DNAase action on bacteriophage T7 DNA showed that endonucleolytic cleavage had occurred by the introduction of single-strand breaks in both strands of the duplex . The molecular weight of the enzyme was determined, by gel filtration on Sephadex G-75, to be 12000.

J Biol Chem, 1978 Apr 10, 253(7), 2238 - 43
Isolation of 4-(1'-D-ribitylamino)-5-amino-2,6-dihydroxypyrimidine from a riboflavin-adenine-deficient mutant of Bacillus subtilis; Mitsuda H et al.; Studies were carried out to determine possible intermediates involved in the biosynthetic pathway of riboflavin, using resting cells of a riboflavin-adenine-deficient mutant, Bacillus subtilis AJ1988 . The cells excreted 6,7-dimethyl-8-ribityllumazine, the end product in the biosynthetic pathway, into the incubation medium in large amounts . The addition of glyoxal caused a large accumulation of a green fluorescent compound; an inverse relation was observed between the formation of the lumazine and the concentration of glyoxal . Furthermore, added {2-14C}guanine effectively incorporated into the lumazine and the fluorescent compound in the same specific activity during incubation . The fluorescent compound was isolated, purified, and identified by paper chromatographic, fluorometric, and spectrophotometric analyses . It was proved to be 8-(1'-D-ribityl)lumazine, which appeared to have been formed by a reaction between glyoxal and a possible intermediate in the cells . Accordingly, 4-(1'-D-ribitylamino)-5-amino-2,6-dihydroxypyrimidine was concluded to be an immediate precursor of 6,7-dimethyl-8-ribityllumazine.

Mol Gen Genet, 1978 Apr 6, 160(2), 131 - 7
The initial attachment of transforming DNA to competent Bacillus subtilis; Weppner WA et al.; The initial attachment of transforming DNA to competent Bacillus subtilis is temperature independent between 25 degrees and 45 degrees . However, below 15 degrees there is a significant reduction in the amount of DNA attached to compentent cells . The DNA that is attached at 4 degrees can lead to transformation or interfere effectively with the subsequent attachment of a distinctive DNA when the cells are shifted to a permissive temperature (37 degrees) . These data suggest that the attachment of DNA at 4 degrees is to sites normally involved in the transformation process . The amount of DNA that is initially attached to the bacteria at 4 degrees or 37 degrees after perturbation of the cells by ionic strength changes, repetitive washings, or periodate oxidation varies with the temperature at which the treatment occurs . These results are consistent with a reorientation of the DNA attachment sites upon lowering the temperature to 4 degrees, such that their affinity for DNA and susceptibility inhibitory treatments are reduced.

J Pharm Sci, 1978 Apr, 67(4), 576 - 8
Synthesis and antimicrobial activity of ethyl N-aryl-S-(triphenylstannyl)isothiocarbamates; Kupchik EJ et al.; Five ethyl N-aryl-S-(triphenylstannyl)isothiocarbamates were synthesized by the reaction of triphenyltin iodide with the appropriate ethyl N-arylthiocarbamate in the presence of triethylamine . The IR spectrum of each compound was obtained over the 4000--200-cm--1 range, and some bands were assigned . These new compounds were found to be generally better antifungal agents than the previously tested N-substituted N'-cyano-S-(triphenylstannyl)isothioureas . The new compounds were also investigated for antibacterial activity and were especially inhibitory toward Gram-positive species . Except for their lower activity toward Bacillus subtilis, their antibacterial activity was identical to the previously tested N-phenyl-N'-cyano-S-(triphenylstannyl)isothiourea.

J Bacteriol, 1978 Apr, 134(1), 318 - 29
Characterization of Staphylococcus aureus plasmids introduced by transformation into Bacillus subtilis; Gryczan TJ et al.; Covalently closed circular DNA from five Staphylococcus aureus plasmids has been introduced into Bacillus subtilis . Four of these plasmids (pUB110, pCM194, pSA2100, and pSA0501) have been selected for further study . These plasmids replicate as multicopy autonomous replicons in both Rec+ and Rec- B . subtilis strains . They may be transduced between B . subtilis strains or transformed at a frequency of 10(4) to 10(5) transformants per microgram of DNA . The molecular weights of these plasmids were estimated, and restriction endonuclease cleavage site maps are presented . Evidence is given that pSA2100, an in vivo recombinant of pSA0501 and pCM194 (S . Iordanescu, J . Bacteriol . 124:597-601, 1975), arose by a fusion of the latter plasmids, possibly by insertion of one element into another as a translocatable element . Genetic information from three other S . aureus plasmids (pK545, pSH2, and pUB101) has also been introduced into B . subtilis, although no covalently closed circular plasmid DNA was recovered.

J Bacteriol, 1978 Apr, 134(1), 24 - 9
Deoxyuridine residues in DNA of thymine-requiring Bacillus subtilis strains with defective N-glycosidase activity for uracil-containing DNA; Makino F et al.; DNA extracted from exponentially growing cells of thymine-requiring Bacillus subtilis strains with defective N-glycosidase activity for deoxyuridine residues in DNA was subjected to the action of N-glycosidase in vitro and analyzed by sedimentation in alkaline sucrose gradients . The sites attacked by N-glycosidase occurred once per 6 X 10(6) to 7 X 10(6) daltons of DNA from cells cultured in the presence of growth-supporting concentrations of thymine . The number of N-glycosidase-susceptible sites increased when the thymine concentration in the medium was lowered . Parallel to this observation, the N-glycosidase-defective mutant cells were less apt to show the detrimental effect due to thymine depletion than were the parental cells . Such sites were not detected in DNA from cells with a normal N-glycosidase activity or with a "wild type" capacity for thymidylate synthesis . The results are interpreted to mean that cells defective for thymidylate synthesis incorporate dUTP in place of TTP in DNA and that the deoxyuridine residues, once incorporated, remain in the DNA in the absence of N-glycosidase activity.

J Gen Microbiol, 1978 Apr, 105(2), 175 - 85
A heat-sensitive lysis mutant of Bacillus subtilis 168 with a low activity of pyruvate carboxylase; Buxton RS; A mutant of Bacillus subtilis which grew in complex medium at 30 degrees C but lysed at 45 degrees C has been isolated . It could only grow on minimal medium at 45 degrees C with added aspartate (20 microgram ml-1) but lysed if lysine (20 microgram ml-1) was also present . The requirement for aspartate was due to a low activity of pyruvate carboxylase; the site of the mutation (pyc) was linked (16% cotransducible using phage PBSI) to the pyrD locus, and the order of markers deduced was: pyrD-cysC-pyc . This defect appeared to lead to decreased synthesis of mesodiaminopimelic acid (mesoA2pm), an amino acid unique to peptidoglycan and its precursors . At the restrictive temperature the mutant accumulated uridine-5'-diphosphate N-acetylmuramyl-L-alanyl-D-glutamate, since meso A2pm is the next amino acid to be added to the growing peptide chain of peptidoglycan . This resulted in an inhibition of peptidoglycan synthesis, determined as a reduced incorporation of N-acetyl{14C}glucosamine . Peptidoglycan synthesis was not decreased if the mutant was grown in media containing aspartate but lacking lysine . The sensitivity to lysine may arise because (i) at 45 degrees C the mutant was starved for aspartate and hence mesoA2pm even when aspartate was present, since aspartate utilization, as estimated by the incorporation of {3H}aspartate into trichloroacetic acid precipitable material, was relatively inefficient; and (ii) this diminished level of mesoA2pm synthesis from aspartate was further curtailed since lysine inhibits one of the aspartokinases in B . subtilis . Thus, addition of lysine allowed protein synthesis and hence autolysin production to proceed whilst peptidoglycan synthesis remained inhibited . When autolysis was blocked, either indirectly by stopping protein synthesis through starvation of aspartate and lysine, or directly by introducing a lyt mutation, then shifting the mutant to 45 degrees C did not result in lysis but growth still ceased.

Int J Radiat Biol Relat Stud Phys Chem Med, 1978 Apr, 33(4), 317 - 24
Radiation sensitivity of transforming DNA; Held KD et al.; Biologically active DNA isolated from Bacillus subtilis was exposed in vitro to X-rays at a concentration of 10 microgram/ml in 29 mM phosphate buffer . Radiation-induced damage to the DNA was quantitatively determined by measuring the decrease in its transforming activity (try2 locus) using B . subtilis 168M (try-) as recipient . In O2, which removes .H and eaq-, the radiation sensitivity of the DNA is less than that in N2-saturated water . In N2O, which has been shown to increase yields of .OH in irardiated aqueous solutions, the radiation sensitivity of Transforming DNA is twice that observed in O2 and 1.5 times that in N2 . Addition of 5 X 10(-2) M ethanol or 1.7 X 10(-1) M t-butanol, both .OH scavengers, causes large (about tenfold) reduction in the radiation sensitivity in all three saturating gases . These results suggest the importance of the .OH radical in the loss of biological activity of DNA.

J Antibiot (Tokyo), 1978 Apr, 31(4), 284 - 8
Identification of antibiotics of iturin group in various strains of Bacillus subtilis; Besson F et al.; Thirty eight strains of B . subtilis were tested for the presence of antifungal antibiotics of the iturin group . The characterization of these antibiotics was made on the basis of: antifungal activity against P . chrysogenum, isolation and purification of the active substance, quantitative analysis of alpha-aminoacids and identification of the liposoluble dipeptide obtained by partial hydrolysis . The only antibiotics of the iturin group found were: iturin A, mycosubtilin and bacillomycin L.

J Bacteriol, 1978 Apr, 134(1), 108 - 14
Alkaline phosphatase possessing alkaline phosphodiesterase activity and other phosphodiesterases in Bacillus subtilis; Yamane K et al.; In Bacillus subtilis Marburg strain, single-point mutations in the phoP locus brought about simultaneous losses of the major activities of alkaline phosphatase (APase) and alkaline phosphodiesterase (APDase) . Revertants recovered the two activities . APases with APDase activity were purified from the membrane fraction of B . subtilis 6160-BC6 and from the culture fluid of an APase-secreting B . subtilis mutant strain, RAN 1 . In addition to these major APases with APDase activity, at least two kinds of phosphodiesterase (PDase) without phosphatase activity were found in the cytoplasmic supernatants of RAN 1 and an APase-less B . subtilis mutant strain, SP25 . Another minor APase with a molecular weight of about 80,000, which had almost no PDase activity, was isolated from the membrane fraction of strain 6160-BC6 . Enzyme distribution in subcellular fractions from various strains cultured in high- and low-phosphate media was analyzed . The PDases did not cross-react with rabbit antiserum against the RAN 1 APase with APDase activity . The main component of the PDases had a molecular weight of about 80,000 and was most active at pH 8.0 . These results suggest that APase with APDase activity is different from PDases detected in cytoplasmic supernatants and that phoP is the structural gene for the phosphate-repressible APase with APDase activity.

J Bacteriol, 1978 Apr, 134(1), 100 - 7
Purification and characterization of extracellular soluble and membrane-bound insoluble alkaline phosphatases possessing phosphodiesterase activities in Bacillus subtilis; Yamane K et al.; A membrane-bound insoluble alkaline phosphatase (APase) and an extracellular soluble APase were purified, respectively, from a membrane preparation of Bacillus subtilis 6160-BC6, which carries a mutation to produce APase constitutively, and from a culture fluid of a mutant strain . RAN 1, isolated from strain 6160-BC6, which produces an extracellular soluble APase . The two preparations were homogeneous, as judged by sodium dodecyl sulfate discontinuous gel electrophoresis and by gel electrophoreses in the presence of 8 M urea at pH 9.3 and 4.3 . RAN 1 APase was crystallized . Both preparations possessed phosphatase and phosphodiesterase activities, and their pH optima were both at 9.5 . They were competitively inhibited by phosphate or arsenate and were activated by the addition of Ca2+ but not by Zn2+ . The APase and alkaline phosphodiesterase activities seemed to be contained in the same protein molecule . The molecular weight of 6160-BC6 APase was estimated to be 46,000 +/- 1,000, and that of RAN 1 APase was estimated to be 45,000 +/- 1,000 . The largest difference between the 6160-BC6 and RAN 1 APase's was in solubility in low-ionic-strength solutions . Present results suggest that each enzyme is composed of a single polypeptide chain and that 6160-BC6 APase aggregates in solutions of low ionic strength.

J Biol Chem, 1978 Mar 25, 253(6), 1756 - 65
Role of the 21,000 molecular weight polypeptide of Bacillus subtilis RNA polymerase in RNA synthesis; Spiegelman GB et al.; DNA-dependent RNA polymerase from Bacillus subtilis contains a 21,000 molecular weight (21K) peptide subunit . This subunit was purified and added to 21K-depleted polymerase isolated from both uninfected and SP85-infected B . subtilis . The effect of the subunit on total RNA synthesis, on enzyme-DNA binding, on RNA chain initiation and elongation, and on enzymatic specificity were examined . A comparison was made of the effects of the 21K peptide and NaCl on polymerase activity, RNA chain elongation, and symmetry of transcription of SP82 DNA . The addition of the 21K peptide to enzyme preparations lacking this subunit stimulated total polymerase activity 20 to 40% . In contrast, addition of NaCl at concentrations greater than 0.1 M significantly reduced polymerase activity . The 21K peptide appeared to alter the general affinity of the polymerase for DNA . The rate of RNA chain initiation was not affected by the 21K peptide, but RNA chain elongation was stimulated . Both the 21K peptide and NaCl increased the asymmetry of transcription of SP82 DNA by phage-modified polymerase . The 21K effect was related to the stimulation of elongation while high concentrations of NCl appeared to act at RNA chain initiation . RNAs synthesized in vitro by polymerase lacking and supplemented with the 21K peptide were translated by a Escherichia coli cell-free system . The 21K peptide had little direct effect on the selection of promoters in vitro as measured by this technique, but it dramatically increased the translatability of the product.

Biochim Biophys Acta, 1978 Mar 20, 539(3), 386 - 97
Effects of staphylococcin 1580 on cells and membrane vesicles of Bacillus subtilis W23; Weerkamp A et al.; 1 . Uptake of L-glutamic acid is inhibited, and preaccumulated L-glutamic acid is released from Bacillus subtilis cells treated with staphylococcin 1580 . Uptake of alpha-methylglycoside is enhanced at low bacteriocin concentrations and inhibited by excess bacteriocin . 2 . Inhibition of amino acid uptake into membrane vesicles is somewhat less sensitive to staphylococcin 1580 than uptake into whole cells under similar conditions, when the bacteriocin concentration is expressed per weight unit of membrane protein . Inhibition of uptake into vesicles is independent of the electron donor system used, but varies with different substrates . 3 . Influx of L-glutamic acid into vesicles under anaerobic conditions is severely hampered by staphylococcin 1580 . The L-glutamic acid carrier functions are slightly affected only . 4 . Staphylococcin 1580 abolished the membrane potential induced by respiration or by a potassium diffusion potential in the presence of valinomycin, as measured with the fluorescent dye 3,3'-dipropylthiadicarbocyanine . 5 . The effects of staphylococcin 1580 on cells and membrane vesicles allowed the classification into three groups with different sensitivity to the staphylococcin concentration.

Eur J Biochem, 1978 Mar 15, 84(2), 533 - 9
Properties of the high-molecular-weight deoxyribonuclease of Bacillus subtilis degrading single-stranded DNA; Cobianchi F et al.; We have studied the properties of the high-Mr DNAse degrading single-stranded DNA which is present in extracts of Bacillus subtilis . This enzyme is a heterogeneous aggregate of identical subunits with an Mr of 36 000, as measured in dodecylsulfate/polyacrylamide electrophoresis . The aggregate can be disassembled by the presence of Triton X-100, but reforms spontaneously following removal of the detergent . A mild proteolytic treatment of the aggregate causes the irreversible and nearly quantitative conversion into the free subunit . The modified subunit has identical properties (in terms of size, chromatographic adsorption and catalytic activity) as the small DNAse previously described by Ciarrocchi et al . {Eur . J . Biochem . 61, 487 (1976)}, i.e . an endonuclease highly specific for single-stranded DNA and producing 5'-P and 3'-OH ends.

Lloydia, 1978 Mar-Apr, 41(2), 156 - 60
Antimicrobial activities of constituents of Uvaria chamae; Hufford CD et al.; The antimicrobial activities of a number of cytotoxic C-benzylated flavonoids from Uvaria chamae have been determined . The minimum inhibitory concentration values of these flavonoids and certain of their derivatives against Straphylococcus aureus, Bacillus subtilis, and Mycobacterium smegmatis compare favorably with those of streptomycin sulfate.

Proc Natl Acad Sci U S A, 1978 Mar, 75(3), 1433 - 6
DNA cloning in Bacillus subtilis; Ehrlich SD; A plasmid pC194, encoding resistance to chloramphenicol, can serve as a cloning vector in Bacillus subtilis 168 for other HindIII-cleaved DNA segments . Replicons constructed by linking pC194 to several Escherichia coli plasmids can be used to introduce and compare the expression of the same genes in these two bacterial hosts.

Proc Natl Acad Sci U S A, 1978 Mar, 75(3), 1423 - 7
Molecular cloning of genetically active fragments of Bacillus DNA in Bacillus subtilis and properties of the vector plasmid pUB110; Keggins KM et al.; Plasmid pUB110 (approximately 2.8 x 10(6) daltons), originally detected in Staphylococcus aureus, specifies resistance to neomycin and has been transformed into Bacillus subtilis 168, In B . subtilis, pUB110 is stably maintained at about 50 copies per chromosome and renders the host resistant to neomycin sulfate at 5 microgram/ml . pUB110 isolated from B . subtilis transforms Rec+ and recE4-containing strains of B . subtilis at frequencies greater than or equal to 10(3) transformants per microgram of plasmid . pUB110 was transferred by PBS1 or SP10 transduction from B . subtilis to strains of B . pumilus and B . licheniformis . pUB110 is compatible with each of four previously described Bacillus plasmids, including pPL576, pPL10, pPL7065, and pPL2 . pUB110 contains a single EcoR1-sensitive site and was used as vector to clone DNA fragments that complement the trpC2 mutation in B . subtilis 168 from EcoR1 digests of the chromosome DNA isolated from B . pumilus strains NRRL B-3275 and NRS576, B . licheniformis strains 9945A and 749C, and B . subtilis 168 . Genetic and physical properties of each of the constructed Trp derivatives of pUB110 are described.

Proc Natl Acad Sci U S A, 1978 Mar, 75(3), 1194 - 8
Two thymidylate synthetases in Bacillus subtilis; Neuhard J et al.; Bacillus subtilis grown at temperatures below 37 degrees contains two thymidylate synthetases, TSaseA and TSaseB . Their presence is dependent on functional thyA and thyB genes, respectively . When cells are grown at 46 degrees they only contain TSaseA activity . This allous an easy positive selection for thyA and thyA, THYB mutants . The two TSases have been physically separated, and they show similar overall requirements for activity . However, they differ significantly in both their kinetic and their physicochemical properties.

Proc Natl Acad Sci U S A, 1978 Mar, 75(3), 1185 - 9
Promoter recognition by phage SP01-modified RNA polymerase; Talkington C et al.; A modified form of Bacillus subtilis RNA polymerase containing a phage SP01-coded regulatory protein (the gene 28 product) selectively transcribes "middle" genes of the phage genome in vitro . In this paper, we identify a subset of restriction endonuclease fragments of SP01 DNA that promote specific transcription by the phage-modified polymerase . In the absence of nucleoside triphosphates, RNA polymerase containing the gene 28 protein selectively binds to these DNA fragments thereby forming stable binary complexes that can be isolated on nitrocellulose filters . In contrast, unmodified RNA polymerase containing sigma factor selectively binds to and transcribes a subset of phage DNA fragments that contain "early" sequences and that are in large part distinct from the fragments recognized by the phage-modified transcriptase . Our results strongly suggest that phage "early" and "middle" genes are transcribed from distinct promoters and that the RNA polymerase containing the gene 28 protein binds to sites that are located at or near promoters for SP01 "middle" genes.

Can J Microbiol, 1978 Mar, 24(3), 282 - 8
Genetic fate of DNA in a strain of Bacillus subtilis which is impaired in genetic transformation; Devanathan T et al.; A mutation in Bacillus subtilis call recC4 which results in an impairment of genetic transformation was transferred to a new strain using the closely linked marker mit-2 (mitomycin C-resistance) for selection . This derived strain was in turn impaired in transformation but showed normal levels of sensitivity to ultraviolet irradiation and methyl methane sulfonate . The genetic and molecular fate of transforming DNA in the recC4 strain was studied . Normal amounts of DNA were taken up by the cells and this DNA or parts of it became associated with recipient DNA . Linkage between genes on donor and recipient molecules was, however, not established and transformants were not generated . The recC4 mutation therefore affects a step in the recombination pathway during transformation . Either the association between donor and recipient DNA molecules is abnormal or the cells are deficient in the further processing of the associated complex.

Arch Microbiol, 1978 Mar, 116(3), 245 - 52
Metabolic products of microorganisms 167 . Cyclopaldic acid from Aspergillus duricaulis . 1 . Production, isolation and bioloical properties; Brillinger GU et al.; In the course of a screening for new metabolites from fungi we isolated a substance with antimicrobial activity from cultures of Aspergillus duricaulis (CBS 481.65) (Tu 679) . It was antagonized by putrescine, spermidine, spermine, arginine, citrulline, lysine, ornithine, in higher concentration by aspraagine and glutamine too . The effect of ethylenediaminetetraacetate on the susceptibility of Streptomyces viridochromogenes (Tu 57) and Bacillus subtilis ATCC 6051 to this antibiotic has been studied . The substance was characterized and identified as cyclopaldic acid.

J Bacteriol, 1978 Mar, 133(3), 1339 - 50
D-alanine incorporation into macromolecules and effects of D-alanine deprivation on active transport in Bacillus subtilis; Clark VL et al.; An auxotroph of Bacillus subtilis 168 unable to synthesize D-alanine loses the ability to support endogenously energized transport when deprived of D-alanine . Revertants of the mutant retain transport activity . The loss of transport is specific for substrates taken up by active transport; substrates taken up by group translocation are transported at normal rates . The loss of transport can be retarded by pretreatment of the cells with inhibitors of protein synthesis . Since the loss of transport could be due to an alteration in a D-alanine-containing polymer, we investigated the incorporation of D-{14C}alanine into macromolecules . The major D-alanine-containing polymers in B . subtilis are peptidoglycan and teichoic acid, with 4 to 6% of the D-{14C}alanine label found in trypsin-soluble material . Whereas the peptidoglycan and teichoic acid undergo turnover, the trypsin-soluble material does not . Treatment of the trypsin-soluble material with Pronase releases free D-alanine . Analysis of acid-hydrolyzed trypsin-soluble material indicated that approximately 75% of the radioactivity is present as D-alanine, with the remainder present as L-alanine . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of partially purified D-{14C}alanine-labeled membranes indicated the presence of two peaks of radioactivity (molecular weights, 230,000 and 80,000) that could be digested by trypsin . The results suggest that D-alanine may be covalently bound to cellular proteins.

J Bacteriol, 1978 Mar, 133(3), 1246 - 53
Interspecies transformation in Bacillus: mechanism of heterologous intergenote transformation; Harris-Warrick RM et al.; Bacillus subtilis-Bacillus globigii hybrids were made by integration of the B . globigii aromatic region (aroB to aroE) as an intergenote in the B . subtillis chromosome . Transformation of the heterologous intergenote by B . subtillis DNA (or vice versa) occurred at about 10% of the frequency of homologous transformation by hybrid donors into the same region . Heterologous intergenote crosses were unusually sensitive to shear fragmentations of donor DNA to sizes less than 30 X 10(6) to 40 X 10(6) daltons . In all cases, the entire intergenote was transferred en bloc . Homologous transformation of intergenote markers by B . globigii DNA was not unusually shear sensitive, and linkage was normal for markers in the intergenote . A model is proposed in which efficient heterologous intergenote transformation occurs by recognition and base pairing of homologous DNA sequences of both flanks of the intergenote.

J Bacteriol, 1978 Mar, 133(3), 1237 - 45
Interspecies transformation in Bacillus: sequence heterology as the major barrier; Harris-Warrick RM et al.; The relative contribution of DNA restriction and of sequence heterology as barriers to interspecies transfer of DNA was studied in the heterologous transformation of Bacillus subtilis recipients by DNA was studied in the heterologous transformation of Bacillus subtilis recipients by DNA isolated from B . globigii . Transformants were obtained at very low frequencies in the evolutionarily nonconserved aromatic region; high cotransfer of linked markers was observed . New mutations were introduced into the B . globigii intergenote sequence in the resulting hybrids; these markers could be transformed with high efficiency by both B . globigii and B . subtilis DNA, representing a 10(5)-fold increase in heterologous transforming efficiency . A restriction activity in B . globigii crude extracts inactivated the biological activity of B . subtilis and hybrid DNA but not B . globigii DNA in vitro, demonstrating different sites for restriction and modification between these species . In vivo, however, B . globigii and hybrid DNA transformed the B . globigii sequence in a hybrid recipient with the same efficiency . These results show that sequence heterology is the major barrier to interspecies transformation and that, in this system, enzymatic restriction does not prevent interspecies transformation.

Can J Biochem, 1978 Mar, 56(3), 158 - 60
Synthesis of thymidine nucleotides in Bacillus subtilis; Rima BK et al.; To investigate the synthesis of thymidine nucleotides in Bacillus subtilis, mutants that carried various combinations of thyA, thyB, and other mutations affecting pyrimidine metabolism were isolated . It was found that exogenously supplied deoxycytidine was converted to thymidine nucleotides . The present data suggest that deoxycytidine nucleotides are first deaminated to yield deoxyuridine nucleotides which can serve as substrates for both thyA- and thyB-coded synthetases . A deaminase activity for dCDP was found in crude extracts of B . subtilis . A mutant lacking the deaminase activity was unable to convert deoxycytidine nucleotides to thymidine nucleotides.

J Bacteriol, 1978 Mar, 133(3), 1401 - 11
Intracellular serine protease of Bacillus subtilis: sequence homology with extracellular subtilisins; Strongin AY et al.; Intracellular serine protease was isolated from stationary-grown Bacillus subtilis A-50 cells and purified to homogeneity . The molecular weight of the enzyme is 31,000 +/- 1,000, with an isoelectric point of 4.3 . Its amino acid composition is characteristically enriched in glutamic acid content, differing from that of extra-cellular subtilisins . The enzyme is completely inhibited with phenylmethylsulfonyl fluoride and ethylenediaminetetraacetic acid . Intracellular protease possesses negligible activity towards bovine serum albumin and hemoglobin, but has 5- to 20-fold higher specific activity against p-nitroanilides of benzyloxycarbonyl tripeptides than subtilisin BPN' . Esterolytic activity of the enzyme is also higher than that of subtilisin BPN' . The enzyme is sequence homologous with secretory subtilisins throughout 50 determined NH2-terminal residues, indicating the presence of duplicated structural genes for serine proteases in the B . subtilis genome . The occurrence of two homologous genes in the cell might accelerate the evolution of serine protease not only by the loosening of selective constrainst, but also by creation of sequence variants by means of intragenic recombination . Three molecular forms of intracellular protease were found, two of them with NH2-terminal glutamic acid and one minor form, three residues longer, with asparagine as NH2 terminus . These data indicate the possible presence of an enzyme precursor proteolytically modified during cell growth.

J Bacteriol, 1978 Mar, 133(3), 1083 - 8
Protonmotive force and motility of Bacillus subtilis; Shioi JI et al.; Motility of Bacillus subtilis was inhibited within a few minutes by a combination of valinomycin and a high concentration of potassium ions in the medium at neutral pH . Motility was restored by lowering the concentration of valinomycin or potassium ions . The valinomycin concentration necessary for motility inhibition was determined at various concentrations of potassium ions and various pH's . At pH 7.5, valinomycin of any concentration did not inhibit the motility, when the potassium ion concentration was lower than 9 mM . In the presence of 230 mM potassium ion, the motility inhibition by valinomycin was not detected at pH lower than 6.1 . These results are easily explained by the idea that the motility of B . subtilis is supported by the electrochemical potential difference of the proton across the membrane, or the protonmotive force . The electrochemical potential difference necessary for motility was estimated to be about -90 mV.

Mol Gen Genet, 1978 Feb 27, 159(3), 337 - 9
Two related structural genes coding two homologous serine proteases in the Bacillus subtilis genome; Strongin AY et al.; A Bacillus subtilis intracellular serine protease is homologous in sequence to an extracellular serine protease from the same strain, indicating the presence of two related structural genes in the Bacillus subtilis genome.

Mol Gen Genet, 1978 Feb 16, 159(2), 213 - 8
Differences in pattern of a DNA protein complex isolated from vegetative cells and spores of Bacillus subtilis; Gruissem W; A DNA protein complex has been isolated from vegetative cells and spores of Bacillus subtilis . Properties of the DNA protein complex prepared from vegetative cells were studied and SDS gel electrophoresis was employed to compare the different DNase-untreated and -treated DNA protein complexes . It is concluded that proteins are associated with the DNA and differences in protein pattern in polyacrylamide gels indicates the involvement of DNA-binding proteins in the regulation of spore formation.

Nucleic Acids Res, 1978 Feb, 5(2), 537 - 48
Nucleotide sequence of threonine tRNA from Bacillus subtilis; Hasegawa T et al.; A threonine tRNA was purified from Bacillus subtilis W168 by a combined use of column chromatographic systems . The nucleotide sequence was determined to be pG-C-C-G-G-U-G-U-A-G-C-U-C-A-A-U-D-G-G-D(U)-A-G-A-G-C-A-A-C-U-G-A-C-U-mo5U-G-U-t6A-A-psi-C-A-G-U-A-G-m7G-U-U-G-G-G-G-G-T-psi-C-A-A-G-U-C-C-U-C-U-U-G-C-C-G-G-C-A-C-C-AOH, where about 40 % of D20 remained unmodified as U20 . It consists of 76 nucleotides including a new minor nucleoside, 5-methoxyuridine (mo5U), which occupies the wobble position of anticodon.

Nucleic Acids Res, 1978 Feb, 5(2), 475 - 89
Folded chromosomes of vegetative Bacillus subtilis: composition and properties; Guillen N et al.; The isolation of folded DNA from Bacillus subtilis, a Gram positive bacterium is described . When the lysis was achieved with 1 M NaCl a slow sedimenting nucleoid was obtained (1600-2000 S) . Conversely, when the lysis was achieved with 0.2 M NaCl a fast sedimenting nucleoid was obtained (3500-4000 S) . The yield of folded DNA was between 80 to 90 % of the total lysate DNA . Both nucleoids contained the same amount of RNA, but the relative proportions of lipids and proteins were different . Folded chromosomes were prepared in the presence of spermidine: artifactual protein binding is shown to be unlikely . Electrophoresis of nucleoid proteins showed a dominant polypeptide (MW = 36,000), which remained associated with DNA after sarcosyl treatment and could be partially removed by heat mediated DNA unfolding . In vitro transcription by endogenous RNA polymerase bound to the fast sedimenting-nucleoid was rifamycin resistant; the template capacity of the fast sedimenting-nucleoid was compared with that of the completely unfolded chromosomes.

J Clin Pathol, 1978 Feb, 31(2), 148 - 52
Sporicidal activity of mixtures of alcohol and hypochlorite; Coates D et al.; In an investigation of the activity against Bacillus subtilis spores of various mixtures of alcohol and sodium hypochlorite 50% methanol with enough hypochlorite to provide 2000 parts per million available chlorine water was the most sporicidal . This mixture, both when freshly prepared and for at least eight hours after mixing, achieved a five-log reduction in spore count in 10 minutes . The sporicidal activity of mixtures containing the same initial level of available chlorine was found to vary with the alcohol, initial alcohol concentration, and age of the mixture . Such mixtures may be useful for disinfecting heat-sensitive instruments.

Mutat Res, 1978 Feb, 49(2), 179 - 86
Mutation induction with UV- and X-radiations in spores and vegetative cells of Bacillus subtilis; Tanooka H et al.; Spores and vegetative cells of Bacillus subtilis strains with various defects in DNA-repair capacities (hcr-, ssp-, hcr-ssp-) were irradiated with UV radiation or X-rays . Induced mutation frequency was determined from the observed frequency of prototrophic reversion of a suppressible auxotrophic mutation . At equal physical dose, after either UV- or X-irradiation, spores were more resistant to mutations as well as to killing than were vegetative cells . However, quantitative comparison revealed that, at equally lethal doses, spores and vegetative cells were almost equally mutable by X-rays whereas spores were considerably less mutable by UV than were vegetative cells . Thus, as judged from their mutagenic efficiency relative to the lethality, X-ray-induced damage in the spore DNA and the vegetative DNA were equally mutagenic, while UV-induced DNA photoproducts in the spore were less mutagenic than those in vegetative cells . Post-treatment of UV-irradiated cells with caffeine decreased the survival and the induced mutation frequency for either spores or vegetative cells for all the strains . In X-irradiated spores, however, a similar suppressing effect of caffeine was observed only for mutability of a strain lacking DNA polymerase I activity.

J Bacteriol, 1978 Feb, 133(2), 897 - 903
Bacteriocin and antibiotic resistance plasmids in Bacillus cereus and Bacillus subtilis; Bernhard K et al.; A number of plasmids have been isolated as covalently closed circular DNAs from strains of Bacillus cereus and B . subtilis . From 12 out of 15 strains of B . cereus, plasmids could be isolated . Most of the B . cereus strains contained two or more plasmids . Their molecular weights ranged from 1.6 X 10(6) to 105 X 10(6) . Bacteriocin production could be attributed to a 45 X 10(6)-dalton plasmid (pBC7) from B . cereus DSM 336, and tetracycline resistance to a 2.8 X 10(6) plasmid (pBC16) from B . cereus GP7 . Two streptomycin-resistant strains of B . subtilis harbored plasmids of 5.2 X 10(6) and 9 X 10(6), respectively, which were, however, not correlated with the antibiotic resistance . The plasmid carrying resistance to tetracycline, pBC16, which was originally isolated from B . cereus, could be subsequently transformed in B . subtilis, where it is stably maintained.

J Bacteriol, 1978 Feb, 133(2), 816 - 21
Functional expression of two Bacillus subtilis chromosomal genes in Escherichia coli; Chi NY et al.; EcoRI-cleaved deoxyribonucleic acid segments carrying two genes from Bacillus subtilis, pyr and leu, have been cloned in Escherichia coli by insertion into a derivative of the E . coli bacteriophage lambda . Lysogenization of pyrimidine- and leucine-requiring auxotrophs of E . coli by the hybrid phages exhibited prototrophic phenotypes, suggesting the expression of B . subtilis genes in E . coli . Upon induction, these lysogens produced lysates capable of transducing E . coli pyr and leu auxotrophs to prototrophy with high frequency . Isolated DNAs of these bacteriophages have the ability to transform B . subtilis auxotrophs to pyr and leu independence and contain EcoRI-cleaved segments which hybridize to corresponding segments of B . subtilis.

J Bacteriol, 1978 Feb, 133(2), 667 - 70
Mapping of a genetic locus that affects glycerol 3-phosphate transport in Bacillus subtilis; Lindgren V; Two types of fosfomycin-resistant mutants of Bacillus subtilis were isolated . Mutants of the first type (GlpT mutants) were resistant to at least 200 microgram of fosfomycin per ml and failed to take up exogenous glycerol 3-phosphate . Mutants of the second type were resistant to lower concentrations of fosfomycin and transported glycerol-3-phosphate as efficiently as wild-type bacteria . The glpT mutations, but not the mutations in the second type of fosfomycin-resistant mutants, map in the cysA-aroI region of the B . subtilis chromosome.

Eur J Biochem, 1978 Feb 1, 83(1), 307 - 11
Purification and characterization of alanine carrier isolated from H-proteins of Bacillus subtilis; Kusaka I et al.; Alanine transport carrier was isolated and purified from H-proteins of Bacillus subtilis . The purified carrier preparation was homogeneous in migration on polyacrylamide gels containing urea or sodium dodecyl sulfate . Electrophoresis on polyacrylamide gels containing dodecyl sulfate showed a single band of molecular weight of about 7500 . 1 mol alanine was bound/mol carrier protein with a dissociation constant of 0.2 micron . The binding was inhibited by p-chloromercuribenzoate and the inhibition was reversed by dithiothreitol.

J Virol, 1978 Feb, 25(2), 500 - 9
Biosynthesis of 5-(4'5'-dihydroxypentyl) uracil as a nucleoside triphosphate in bacteriophage SP15-infected Bacillus subtilis; Walker MS et al.; The nucleoside triphosphate of 5-(4',5'-dihydroxypentyl)uracil (DHPU) was detected in the acid-soluble extract from bacteriophage SP15-infected Bacillus subtilis W23 . No uracil was found in the DNA of either replicating or mature phage . Labeled thymidine added during phage DNA synthesis was incorporated into phage DNA . The presence of DHPU as a nucleoside triphosphate in the acid-soluble pool and the incorporation of thymidine into phage DNA suggest that both DHPU and thymine are incorporated into SP15 DNA via their nucleoside triphosphates . 5-Fluorodeoxyuridine inhibited biosynthesis of SP15 DNA, and this inhibition was reversed by thymidine, resulting in the synthesis of a DNA containing reduced amounts of fully modified DHPU . It is proposed that 5-fluorodeoxyuridine, or its metabolic product, inhibits a step in the biosynthetic pathway to the nucleoside triphosphate of DHPU.

Can J Microbiol, 1978 Feb, 24(2), 89 - 104
The response of cell walls of Bacillus subtilis to metals and to electron-microscopic stains; Beveridge TJ; Purified cell walls of Bacillus subtilis were subjected to solutions of 40 independent metals and the metal uptake, the electron-scattering power of thin sections, and the type of staining response evaluated . This was repeated for six typical electron-microscopic stains (uranyl acetate, uranyl magnesium acetate, osmium tetroxide, Os-meth, osmium-dimethylethylenediamine, and ruthenium red) and one new staining reagent (a potassium platinum chloride - dimethylsulfoxide complex) whose specificity is for amine functions . The reaction of select metals can be specific in terms of both uptake and staining response . Of the metals studied most transition elements had a high affinity for the wall fabric and some (i.e., Sc III, most lanthanides, UIV, ZrIV,HfIV, Fe III, Pd II, Ru III, and In III) may be suitable as contrasting agents for electron microscopy . Furthermore, when the thickness of metal-reacted walls was compared to freeze-each and ultracryotomy data, statistical-dimensional differences were commonly seen, which indicates that wall ultrastructure can be profoundly affected by the type of metal and (or) staining reagent.

Arch Microbiol, 1978 Jan 23, 116(1), 1 - 8
Control of the chemotactic behavior of Bacillus subtilis cells; de Jong MH et al.; The effects of nigericin, valinomycin and some lipophilic cations on the motile behavior of non-starved and methionine-straved Bacillus subtilis cells were studied . For valinomycin and nigericin a quantitative relationship between the flux in the proton-motive force and the duration of the twiddle response was found . Lipophilic cations bind to the ion gate controlling the twiddle frequency and thereby cause the cells to swim smoothly . To explain the transmission of the chemotactic signal a model is given in which receptors, a hyperpolarizing wave, an ion gate and two methylation sites, viz . methyl-accepting chemotaxis proteins and a further methylation site (MT), play a role . For the transmission of the signal caused by an attractant both the hyperpolarizing wave and an interaction between receptor and methylation site (MT) are needed . The methyl-accepting chemotaxis proteins are involved in the adaptation/deadaptation to altered levels of attractant . Artificial changes in the proton-motive force act directly on the ion gate, which finally controlls the twiddle frequency of the cells.

Mol Gen Genet, 1978 Jan 17, 158(3), 263 - 70
Cloning of Bacillus subtilis leucina A, B and C genes with Escherichia coli plasmids and expression of the leuC gene in E . coli; Nagahari K et al.; The leucine genes of Bacillus subtilis have been cloned directly from the chromosomal DNA into Escherichia coli leuB cells by selection for the Leu+ phenotype using RSF2124 as a vector plasmid . The hybrid plasmid designated RSF2124-B.leu contained a 4.2 megadalton fragment derived from B . subtilis DNA, including the leu genes . The fragment had one site susceptible to EcoRI* and another site susceptible to BamNI endonuclease . Among the three fragments produced by EcoRI* and BamNI endonucleases, the 1.2 megadalton fragment had the ability to transform B . subtilis leuA, leuB and leuC auxotrophs to leu+ . However, B . subtilis ilvB and ilvc auxotrophs were not rescued even by the whole 4.2 megadalton fragment present in the hybrid plasmid . beta-Isopropylmalate dehydrogenase (leuB gene product) activity found in E . coli cells containing the hybrid plasmid was about 60% of that in E . coli wild type cells, despite the high copy number (7.8) of the plasmid per chromosome observed.

Mikrobiologiia, 1978 Jan-Feb, 47(1), 37 - 40
{Changes in the intracellular desoxyribonuclease activity in the process of Bacillus subtilis cell growth and division}; Reshetnik OA et al.; DNA synthesis and cell division were studied in Bacillus subtilis in relation to their intracellular DNAase activity . The specific activity of DNAases was found to increase prior to the beginning of DNA synthesis and cell division . Active DNA synthesis and cell division were accompanied with a decrease in the specific activity of intracellular DNAases.

J Biochem (Tokyo), 1978 Jan, 83(1), 19 - 25
Catabolic conversion of sepiapterin to 6-(1-carboxyethoxy)pterin by Bacillus subtilis; Matsuura S et al.; Structural elucidation of an intensely blue fluorescent compound (A) formed from sepiapterin by Bacillus subtilis is described . The structure of the catabolite (A) was found to be 2-amino-6-(1-carboxyethoxy)-4(3H)-pteridinone (9) from both spectroscopic and degradation studies . This was confirmed by an unambiguous synthesis of 9 . The stereochemical structure of the side chain at the 6-position of A was confirmed to be the L(or S) configuration, as in sepiapterin, by analysis of the lactic acid formed from A on acid hydrolysis . This suggests that the side chain is rearranged intact during the catabolic conversion of sepiapterin . A possible mechanism for the conversion is discussed.

Genetika, 1978, 14(1), 171 - 4
{Bacillus subtilis mutant with altered ability for heterologous transformation}; Prozorov AA et al.; Mutant strain of Bacillus subtilis, which produced in certain conditions significantly reduced quantity of trnasformants during transformation by homologous DNA, as compared with transformation by heterologous DNA from Bac . aterrimus, is isolated . The ability to transfection by phage SPO1 DNA and the efficiency of infection of the mutant by this phage are also decreased . The causes of such alterated properties are discussed.

J Virol, 1978 Jan, 25(1), 224 - 37
Transcriptional specificity of a multisubunit RNA polymerase induced by Bacillus subtilis bacteriophage PBS2; Clark S; Bacillus subtilis phage PBS2 induced the synthesis of two temporally defined categories of phage-specified transcripts . The transcription of phage "early" genes was induced almost immediately after infection; this RNA synthesis did not require phage protein synthesis . Phage "late" gene transcription, on the other hand, was induced at an intermediate time in the lytic cycle; this RNA synthesis required the production of phage proteins . Both classes of transcription were resistant to the drug rifampin and, therefore, apparently did not require the rifampin-sensitive component of the host RNA polymerase . A rifampin-resistant, DNA-dependent RNA polymerase was purified from bacteria infected with PBS2 . The highly purified phage polymerase consisted of five distinct subunits that remained associated during zone centrifugation, isoelectric focusing, and disc gel electrophoresis . The synthesis of each of the five polypeptides was induced at an intermediate time in the phage lytic cycle . As judged by hybridization competition, hybridization to DNA restriction fragments, and RNA-RNA annealing, the phage-induced RNA polymerase preferentially and asymmetrically transcribed PBS2 late genes in vitro . These findings suggest that the PBS2 RNA polymerase controls the expression of phage genes late in the lytic cycle.

J Bacteriol, 1978 Jan, 133(1), 298 - 305
Temperature-sensitive nature of the rodB maturation in Bacillus subtilis; Rogers HJ et al.; The Mg2+ requirement of a morphological mutant of Bacillus subtlis, rodB strain 104 was highly temperature sensitive in the presence of halide or nitrate anions . Likewise the morphological change from rod shapes to spheres was dependent upon temperature, the same anions, and the Mg2+ concentration . The three factors interacted . Other rodB mutants behaved similarly . If the rodB strain 104 in its rod form was treated at high temperatures in the absence of either protein or peptidoglycan synthesis and restored to lower temperatures with the syntheses restarted, a partial temporary change toward cocci occurred . In the absence of halides or in the presence of Cl- but not Br-, the cells increased in volume when they changed from rods to cocci.

J Bacteriol, 1978 Jan, 133(1), 265 - 9
Genetics of the alpha-ketoglutarate dehydrogenase complex of Bacillus subtilis; Hoch JA et al.; The enzymatic defects in a number of Bacillus subtilis mutants of the alpha-ketoglutarate dehydrogenase complex lacking activity have been investigated . Mutants in the citK locus, as well as a series of deletions of unknown length covering the citK locus, are deficient in E1 of the complex, alpha-ketoglutarate dehydrogenase, but have normal activities of E2, dehydrolipoyl transsuccinylase, and E3, lipoamide dehydrogenase . The citK mutants and the citL22 mutant show in vitro complementation of alpha-ketoglutarate dehydrogenase complex activity . The citL22 mutant is severely deficient in lipoamide dehydrogenase activity, and, as a result, lacks activity for both the alpha-ketoglutarate and the pyruvate dehydrogenase complexes . Thus, the E3 components of both complexes are identical . The citL22 mutation maps between ura and metC on the chromosome.

J Environ Sci Health C, 1978, 13(1), 1 - 31
Cell binding and growth inhibition by hexachlorophene or decanoate and their reversibility; Levin BC et al.; More than 80% of the hexachlorophene added to a Bacillus subtilis culture binds to the cells . Complete growth inhibition requires 6 x 10(5) molecules bound per cell . In contrast, more than 99% decanoate remains in solution and 3.8 x 10(7) molecules bound per cell are needed to inhibit growth . Centrifugation and resuspension of cells in growth medium removes only decanoate, whereas the addition of 1% bovine serum albumin to the growth medium removes both inhibitors from their binding sites on the cells . The addition of untreated cells to a hexachlorophene-treated culture enables the hexachlorophene molecules to redistribute among all the cells with the result that the inhibited cells can resume growth.

Arzneimittelforschung, 1978, 28(7), 1067 - 9
The chemical composition and stability of bacillomycin S; Esterhuizen B et al.; Bacillomycin S, an antifungal antibiotic from culture filtrates of Bacillus subtilis, was shown to consist of proteinaceous matter (83.6 g/100 g) and lipid material (16 g/100 g) . The amino acid and fatty acid components were determined qualitatively and quantitatively . It was shown that bacillomycin S is stable at physiological pH and it is suggested that its antifungal activity resides in the intact molecule.

Can J Microbiol, 1978 Jan, 24(1), 1 - 8
Adsorption of Bacillus subtilis bacteriophage PBS 1; Wilson JJ et al.; Bacteriophage PBS 1 adsorbs initially on the flagella of its host, Bacillus subtilis (stage I) . The phage can adsorb to both active and inactive flagella . Flagellar attachment is nonspecific as PBS 1 was shown to attach to the flagella of Bacillus species other than the normal host B . subtilis . The phage particle then quickly moves down the length of the flagellum to its base, the final adsorption site . Flagellar motion is required for flagellar base attachment (stage II) . After proper attachment at the flagellar base, the phage tail sheath contracts sending the tail core through the final adsorption site (stage III) . The phage DNA is then injected at this site (stage IV) . Stage I adsorption does not cause loss of motility in PBS 1 -- resistant bacilli . The loss of motility observed upon infection of sensitive cells by PBS 1 may be associated with either stage II or stage III of adsorption.

Biochimie, 1978, 60(11-12), 1283 - 7
The phosphoenolpyruvate : methyl-alpha-d-glucoside phosphotransferase system in Bacillus subtilis Marburg : kinetic studies of enzyme ii and evidence for a phosphoryl enzyme ii intermediate; Marquet M et al.; The Enzyme II complex catalyzing the phosphoryl transfer from P-HPr to sugar in the inducible methyl-alpha-D-glucoside : phosphotransferase system in Bacillus subtilis acts according to a ping-pong mechanism, implying a phosphorylated Enzyme II intermediate . This result is supported by the demonstration of a specific transphosphorylation between {14C} alphaMG and glucose-6-phosphate in the presence of an induced Enzyme II preparation.

Zentralbl Bakteriol Naturwiss, 1978, 133(7-8), 690 - 7
Further studies on the competence development in exponentially growing cultures of Bacillus subtilis; Lopez P et al.; Bacillus subtilis, growing in Bott and Wilson's medium, develops two peaks of competence in batch cultures . The first maximum developed during the exponential growth phase and it has been studied comparatively with the competence level reached in continuous culture at Dt = 2.5 h . In both cases, 100 microgram/ml of arginine inhibited competence . Continuous cultures treated with arginine recovered competence specifically after the addition of Mn2+ . In addition, a reduction in the synthesis of aconitase and fumarase was observed in the arginine-inhibited continuous cultures.

Genetika, 1978, 14(12), 2102 - 12
{Ontogenetic switchover in Bacillus subtilis . II . The dynamics of the stationary phase processes during growth under conditions of catabolite repression}; Rubikas IP et al.; To reveal the pecularities of the growth under the conditions of catabolite repression (medium 2) of Bacillus subtilis and the mutants obtained, the investigations of dynamics of the following processes were carried out: alteration of the pH of the culture exhaustion of glucose in the medium, appearance of the activity of both aconitase in the cells and extracellular metal- and serine proteases in the supernatant, and the appearance of the thermoresistant spores . The following features were observed during the growth under the conditions of catabolite repression: 1 . Bacillus subtilis WB 746 and cgs mutants: the death of the main part of the culture after the Iogarithmic phase of growth (LPG), the presence of the secondary LPG of the survived cells which have the increasing activity of aconitase, the appearance and sharp increase in the extracellular serine protease activity 6 hours before thermoresistant spore formation . In the case of cgs mutants the activity of metal proteases appears and increases during the secondary LPG; 2 . In the culture of cgl mutants the pH is lowered to 5.1 at the end of the LPG and after the glucose exhaustion the death of almost all the culture follows; 3 . cgr mutants: a comparatively high activity of aconitase in the cells is found by the time of the early LPG, and at the end of the LPG the activity of both metal- and serine proteases appear in the supernatant of the culture and the secondary induction of the serine protease activity 6 hours before thermoresistant spore formation is observed . The serine protease activity found in the supernatant before and after the secondary induction of the enzyme belongs to the identical protein . During the stationary phase of the growth of cgr mutants, the high rate of 3H-uridine incorporation into the RNA molecules which have the electrophoretic mobility of mRNA was observed . The sporulation of Bac . subtilis strains under investigation, except cgl mutants, occurs when the culture has reached the definite state: the alkaline pH, the presence of the aconitase activity in the cells and the induced activity of serine protease.

Genetika, 1978, 14(12), 2082 - 90
{Operon of riboflavin biosynthesis in Bacillus subtilis . XV . A study of mutants related to the initial stages of biosynthesis . The origin of the ribityl chain of the riboflavin molecule}; Bresler SE et al.; The incorporation of 14C-labelled guanosine and xanthosine into riboflavin was studied . It is concluded that the ribose mojety of guanosine is converted to the ribityl side chain of riboflavin . Thus the immediate precursor of riboflavin biosynthesis is a guanosine compound . Two classes of the riboflavin-dependent mutants of Bacillus subtilis were studied . They are closely linked to the lysine markers and probably correspond to the initial steps of riboflavin biosynthesis pathway.

Zentralbl Bakteriol Naturwiss, 1978, 133(2), 121 - 4
Production and chemical studies of protease from Bacillus subtilis (SH-6) Egyptian strain; Mahmoud SA et al.; Bacillus subtilis (SH-6) Egyptian strain, isolated from hides, gave the highest protease activity . Luxurious growth and protease production were obtained by the use of a medium containing 8% of potato starch, 0,1 M of ammonium phosphate as carbon and nitrogen sources . Results indicate that borate buffer exerted a deleterious effect on the protease production . Comparing citrate and phosphate buffers, it was found that citrate gave lower protease activity than phosphate . There is a positive response to higher concentrations of phosphate ions . From the above-mentioned medium protease was precipitated and purified . The dried preparation of the enzyme was tested for its chemical composition . It revealed the absence of residual carbohydrate . Tests for phosphorus, sulfur, ferric, zinc, manganese, magnesium, and calcium ions were positive . Amino acids present were: L-leucine, cysteine, Dl-alanine, L-arginine, L-tyrosine, L-aspargine, L-proline, glycine, Dl-valine, L-histidine, L-glutamic, L-lysine, L-aspartic, Dl-tryptophan, L-cystine, Dl-serine and Dl-phenylalanine . Quantitative analysis of the preparation was 0.52% of ash and 14% of nitrogen.

Rev Asoc Argent Microbiol, 1978 Jan-Apr, 10(1), 14 - 19
{Production of alkaline protease from acid cheese whey}; Massucco AE et al.; The use of acid cheese whey as an essential substrate for the production of alkaline protease was studied by growing Bacillus subtilis NRRL 3411 on different conditions of aereation . The maximum enzymatic activity of 17 000 proteolytic units was obtained in the following conditions: 200 RPM, air flow of 1 v/v minutes (KLa 37 h-1), separate sterilization of the medium components and periodical additions of peptone.

Folia Microbiol (Praha), 1978, 23(3), 183 - 93
Non-transformable mutants of Bacillus subtilis defective in the penetration of DNA into the cell; Tichy P et al.; Four non-transformable mutants of Bacillus subtilis 168 defective in the penetration of DNA into the recipient cell were isolated . All mutants were fully non-transformable with mutation in genes influencing irreversible binding of the donor DNA by the recipient cell.

Genetika, 1978, 14(7), 1175 - 84
{Nature of the mutations that determine the ability of Bacillus subtilis A-50 to sporulate at high glucose concentrations in the medium}; Dobrzhanaskaia EO et al.; The ability of Bacillus subtilis A-50 to sporulate in the medium containing high glucose concentrations is caused by at least two mutation types: pts mutations and cat (or tgl) mutations, both of them affecting differently the level of alkaline proteinase synthesis . The decrease of the level of enzyme activity in the case of pts mutation (gluR3 mutant) occurs at the expense of glucose transport disturbance . The mutation cat (tgl) (mutant gluR5) causes the increase in enzyme synthesis at the expense of catabolic resistance to glucose of genes controlling alkaline proteinase synthesis and the spore formation in Bac . subtilis A-50 . cat5(gluR5) and pts3(gluR3) mutations are located on the chromosome of Bac . subtilis in the region metD and argC respectively . The over-synthesis of alkaline proteinase characteristic of Bac . subtilis A-50 is controlled by the polygenic system, as the level of alkaline proteinase synthesis in argA+ transformants makes up 25% of the level of activity of the original strain . The productivity of Bac . subtilis A-50 can be enhanced by introducing an additional cat mutation.

Vopr Med Khim, 1978 Jan-Feb, 24(1), 17 - 22
{Isolation of cationic proteins from rabbit thrombocytes and study of their bacteriostatic activity}; Goriukhina OA et al.; Cation-containing proteins, which possessed the bacteriostatic activity against Grampositive microorganism Bacillus subtilis SHGW, were isolated from rabbit thrombocytes by means of selective extraction . These substances were of heterogenous nature (as shown by polyacrylamide gel disc electrophoresis), including two groups of proteins with molecular weight about 8=10-10(3) and 20=30-10(3) daltons . The bacteriostatic activity of each group of these proteins was studied after gel filtration of the total preparation on Sephadex G=75 . The group of proteins with molecular weight 8=10-10(3) daltons was responsible for bacteriostatic activity of the total cation-containing protein from thrombocytes.

J Environ Sci Health C, 1978, 13(1), 1 - 31
Cell binding and growth inhibition by hexachlorophene of decanoate and their reversibility; Levin BC et al.; More than 80% of the hexachlorophene added to a Bacillus subtilis culture binds to the cells . Complete growth inhibition requires 6 x 10(5) molecules bound per cell . In contrast, more than 99% decanoate remains in solution and 3.8 x 10(7) molecules bound per cell are needed to inhibit growth . Centrifugation and resuspension of cells in growth medium removes only decanoate, whereas the addition of 1% bovine serum albumin to the growth medium removes both inhibitors from their binding sites on the cells . The addition of untreated cells to a hexachlorophene-treated culture enables the hexachlorophene molecules to redistribute among all the cells with the result that the inhibited cells can resume growth.

Acta Microbiol Pol, 1978, 27(3), 225 - 36
Polypeptide antibiotic 26a from Bacillus subtilis . II . Isolation and purification procedures; Jarosz J; Both the analytical and preparative methods by which the preparations of 26a bydrochloride salt with a high antibacterial activity and 20--30% recovery have been obtained from the fermentation fluids of Bacillus subtilis are presented . On an industrial scale the antibiotic can be yielded by absorption of bioactivity on Amberlite CG-50I column and precipitation with picric acid of crude substance from active elutes as adduct which was divided on equilibrated CM--cellulose and finally purified by gel filtration on Sephadex G-25 column . The purified preparation gave a single band by SDS-polyacrylamide gel electrophoresis and one ninhydrin-positive spot by thin-layer chromatography on silica gel G plates corresponding to single zones of bioactivity on bioautograms, and moved as a single peak of almost constant antibacterial activity on Sephadex G-75, G-100 and G-200 columns . The potency of the purest preparations, lot Sephadex G-25, was 6,500--7,000 arbitrary units/mg, and were characterized as follows: purification factor, 57; purity of 98--100% by densitometer scans of SDS-polyacrylamide gels; MIC for Sarcina lutea by twofold agar dilution assay, 0.306 microgram/ml.

Acta Microbiol Pol, 1978, 27(3), 213 - 24
Polypeptide antibiotic 26a from Bacillus subtilis . I . Taxonomy and fermentative production; Jarosz J; In surface cultures on NK/2-Sym's medium, the isolate No . 26a of Bacillus subtilis from the intestinal tract of Galleria mellonella larvae produced three antibacterial substances which were separated by gel filtration on Sephadex G-25 column . The major bioactive compound named 26a had a close resemblance to bacitracin family of polypeptide antibiotics . Two minor active compounds, i.e . a bacteriolytic enzyme with endo-beta-N-acetylmuramidglycanohydrolase (EC . 3 . 2 . 1 . 17) activity and other unidentified factor were usually synthetized in trace amounts . Maximum yield of 26a generally occurred after 120 hour incubation, when the producer reached the stationary growth phase and general sporulation of the bacterial cultures was found . The basal medium of NK/2-Sym supplemented by addition of manganese ions (10(-4) M), d-glucose (1%) and inorganic nitrogen beneficially resulted in antibiotic potency of the fermentation broth . The antibiotics produced by other isolates (Nos 5AK, 15 and 92) have been also analyzed and from their properties they can be tentatively classified as members of bacitracin group polypeptides . A possible role of the antibiotics produced by intestinal Bacillus spp in the formation process of typical gut microflora of G . mellonella is discussed.

Antonie Van Leeuwenhoek, 1978, 44(3-4), 329 - 39
Properties of beta-glucan synthetase from Saccharomyces cerevisiae; Lopez-Romero E et al.; Properties of beta-glucan synthetase from S . cerevisiae were studied . The enzyme exhibited optimal activity at pH 6.7 and 24 C . Km for UDP-glucose was 0.12 mM . Addition of Mg++ or Mn++ stimulated its activity by 60% and 21% respectively . High concentrations of EDTA and hydroxyquinoline were inhibitory . Glucan synthetase was fully active in cell-free extracts . Small concentrations of trypsin or subtilopeptidase A from Bacillus subtilis, caused only a slight increase in glucosyl transferase activity, but larger concentrations destroyed beta-glucan synthetase . Acid proteases were neither stimulatory nor destructive . Thus it seems unlikely that beta-glucan synthetase exists in a zymogen form . Glucan synthetase was unstable . It was inactivated more rapidly at 28 C than at 0 C . The presence of substrate, beta-glucan or the protease inhibitors PMSF, Antipain or Pepstatin A did not protect beta-glucan synthetase from inactivation . Glucan synthetase was not stimulated by addition of cellobiose or beta-glucans . The synthesis of beta-glucans was competitively inhibited by UDP (Ki = 0.45 mM) . Glucono-delta-lactone, a known inhibitor of beta-glucosidases was a strong non-competitive inhibitor of beta-glucan synthetase.

Z Allg Mikrobiol, 1978, 18(4), 243 - 54
Production of valine by a Bacillus sp; Chattopadhyay SP et al.; A bacterium isolated from Burdwan (India) soil was found to accumulate L-valine in the growth medium and identified to be a strain of Bacillus subtilis . The strain is able to grow and accumulate valine in a purely synthetic medium, but supplementation of the synthetic medium with either Casamino acids or yeast extract or with both, significantly improves the yield . The entire fermentation period can be divided into a growth phase and a production phase, which can be prolonged by adjustment of pH to the neutral range . Among the different carbon and nitrogen sources tested glucose at 8.5% and L-glutamic acid at 0.8%, respectively, were found most suitable . Cane sugar molasses tested as a substitute for glucose significantly stimulated growth but valine production was less . Different vitamins tested stimulated growth and valine yield and an inoculum level of 10% (v/v) of the medium was found to be optimal . The yield of valine under optimal conditions was found to be 4.53 g per litre of the medium . Valine has been isolated in crystalline form from the fermented broth by ion exchange resin chromatography and found to be a pure sample of the L-isomer.

Mol Gen Genet, 1977 Dec 30, 158(2), 193 - 200
Streptomycin-resistant, asporogenous mutant of Bacillus subtilis; Campbell KM et al.; A streptomycin-resistant mutantant of Bacillus subtilis that is also asporogenous, was isolated and partially characterized . This strain, SRB15, sporulated at a frequency of about 1% compared to the wild type frequency of greater than 70% . The two phenotypes were inseparable by transformation, suggesting that this strain carries a single mutation that causes it to be both streptomycin-resistant and spore-minus . The mutation cotransduces with cysA, the closest auxotrophic marker to the "ribosomal region" of the B . subtilis chromosome, with a frequency of 68% . SRB15 showed no cross resistence to other antiobiotics tested, including the aminoglycosides kanamycin, neomycin and spectinomycin . Ribosomes obtained from the mutant were at least 200-fold more resistant in vitro to streptomycin than were wild type ribosomes in the translation of phage SPO1 RNA . The kinetics of in vitro translation of this natural message were indistinguishable for mutant and wild type ribosomes . The level of misreading, as measured by poly(U)-directed isoleucine incorporation, by mutant ribosomes was less than that by wild type ribosomes.

J Biol Chem, 1977 Dec 25, 252(24), 9024 - 31
Reconstitution studies show that rifampicin resistance is determined by the largest polypeptide of Bacillus subtilis RNA polymerase; Halling SM et al.; A procedure has been developed to separate the subunits of Bacillus subtilis RNA polymerase rapidly and in good yield . The method involved the use of a blue dextran-Sepharose column which bound the beta' subunit . A phosphocellulose column was used to separate the alpha and beta subunits . During purification, the enzyme eluted from the DNA-cellulose column in three separate forms in the order alpha2betabeta'deltaomega1,alpha2betabeta'omega1, and alpha2betabeta'omega1sigma . Subunit reconstitution studies with RNA polymerase subunits from wild type and a rifampicin-resistant mutant indicated that the largest polypeptide was responsible for rifampicin resistance . Thus, this subunit is referred to as beta . The mobility of the subunits in sodium dodecyl sulfate-polyacrylamide gel electrophoresis cannot be used as the sole criterion for designating the functions of the subunits of RNA polymerase.

Biochim Biophys Acta, 1977 Dec 8, 485(2), 367 - 78
Effects of carrier morphology and buffer diffusion on the expression of enzymatic activity; Konecny J et al.; A very stable esterase (EC 3.1.1.-), which hydrolyses ethyl acetate, cephalosporin C and other acetyl esters with a maximum turnover number of 3-10(2) s-1, was isolated from Bacillus subtilis ATCC 6633 and immobilized on two supports: controlled-pore glass and powdered brick, a representative of carriers having a wide pore-size distribution . Carrier morphology determines diffusion rates and the expression of activity . Rate-limiting mass transfer of buffer leads to apparent losses of activity, gross distortions of molecular pH vs . activity profiles and to apparent deviations from Michaelis-Menten kinetics.

Biochim Biophys Acta, 1977 Dec 2, 479(3), 367 - 9
Purification of cohesive-end-producing restriction endonuclease from Bacillus subtilis G; Hoshino T et al.; A new restriction endonuclease was partially purified from Bacillus subtilis G (IAM1247) . This restriction endonuclease (endonuclease RBsuG) seems to produce cohesive ends at its cleavage site.

J Biochem (Tokyo), 1977 Dec, 82(6), 1567 - 73
Action of T4 endonuclease V on DNA containing various photoproducts; Makino F et al.; The action of T4 endonuclease V on DNA containing various photoproducts was investigated . (1) The enzyme introduced strand breaks in DNA from ultraviolet-irradiated vegetative cells of Bacillus subtilis but not in DNA from irradiated spores of the same organism . DNA irradiated with long wavelength (360 nm peak) ultraviolet light in the presence of 4,5',8-trimethylpsoralen was not attacked by the enzyme . These results indicate that 5-thyminyl 5,6-dihydrothymine (spore photoproduct) and psoralen mediated cross-links in DNA are not recognized by T4 endonuclease V . (2) DNA of phage PBS1, containing uracil in place of thymine, and DNA of phage SPO1, containing hydroxymethyluracil in place of thymine, were fragmented by the enzyme when the DNA's had been irradiated with ultraviolet light . T4 endonuclease V seems to act on DNA with pyrimidine dimers whether the dimers contain thymine residues or not.

Nucleic Acids Res, 1977 Dec, 4(12), 4291 - 303
Nucleotide sequence of a lysine tRNA from Bacillus subtilis; Yamada Y et al.; A lysine tRNA (tRNA1Lys) was purified from Bacillus subtilis W168 by a consecutive use of several column chromatographic systems . The nucleotide sequence was determined to be pG-A-G-C-C-A-U-U-A-G-C-U-C-A-G-U-D-G-G-D-A-G-A-G-C-A-U-C-U-G-A-C-U-U(U*)-U-U-K-A-psi-C-A-G-A-G-G-m7G(G)-U-C-G-A-A-G-G-T-psi-C-G-A-G-U-C-C-U-U-C-A-U-G-G-C-U-C-A-C-C-AOH, where K and U* are unidentified nucleosides . The nucleosides of U34 and m7G46 were partially substituted with U* and G, respectively . The binding ability of lysyl-tRNA1Lys to Escherichia coli ribosomes was stimulated with ApApA as well as ApApG.

J Gen Microbiol, 1977 Dec, 103(2), 359 - 66
Biosynthesis of thiamin in Bacillus subtilis . Isolation of mutants accumulating 4-amino-5-hydroxymethyl-2-methylpyrimidine phosphate; Walter W et al.; Thiamin-deficient mutants of Bacillus subtilis were characterized by their growth responses to the pyrimidine and thiazole moieties of the vitamin molecule and by cross-feeding tests . All mutants growing on the thiazole moiety and all mutants with an absolute requirement for thiamin fed all those growing on the pyrimidine moiety . No other cross-feeding effects were observed . From the culture fluid of a mutant growing on the thiazole moiety, two compounds were isolated which supported growth of mutants requiring the pyrimidine moiety . These compounds were identified by chromatographic, bioautographic and spectrophotometric procedures as 4-amino-5-hydroxymethyl-2-methylpyrimidine and its monophosphate derivative.

J Bacteriol, 1977 Dec, 132(3), 856 - 61
Repair and subsequent fragmentation of deoxyribonucleic acid in ultraviolet-irradiated Bacillus subtilis recA; Hadden CT; Cells of Bacillus subtilis recA1 are sensitive to irradiation with ultraviolet light . Evidence is presented here that these cells are not defective in ultraviolet light-induced incision of deoxyribonucleic acid (DNA) or repair DNA synthesis . Ligation of DNA at repair sites appears to occur, but the DNA is subsequently fragmented, apparently at sites of previous repair synthesis . It is hypothesized that the defect in DNA repair leads to host-specific restriction at repaired sites because of a defect in either the structure of the repaired region or specificity of the restriction/modification system.

J Bacteriol, 1977 Dec, 132(3), 847 - 55
Restriction-like phenomena in transformation of Bacillus subtilis recA; Hadden CT; Genetic transformation in recA1 strains of Bacillus subtilis was studied to test the hypothesis that, in these strains, a major pathway of recombination is missing, leaving only residual transformation via a pathway specific for transduction . The two putative recombinational pathways have been hypothesized to differ in either length of synapsed regions or specificity for nucleotide sequence homology . It was found that the efficiency of transformation of recA1 cells by deoxyribonucleic acid (DNA) from the heterologous strain W23 was much lower than when a homologous donor DNA was used, the relative efficiency being different for different genetic markers . Because the frequency of recombination between linked markers is only slightly changed in recA1 recipients, and because markers of heterologous origin in DNA from intergenotic strains are not discriminated against strongly by recA1 recipients, it is concluded that neither a difference in length of synapsed DNA nor a difference in specificity for nucleotide sequence homology accounts for reduced transformation in recA1 cells . It is proposed that at some time between uptake and integration, heterologous DNA is inactivated by restriction, and that aberrant restriction of repaired regions may account for reduced transformation by homologous DNA.

Appl Environ Microbiol, 1977 Dec, 34(6), 689 - 94
Increase in arginine and citrulline production by 6-azauracil-resistant mutants of Bacillus subtilis; Kato J et al.; In the arginine producer AHr-5, an L-arginine hydroxamate-resistant mutant of Bacillus subtilis, accumulation of N8-acetyl-L-ornithine increased as the level of L-arginine accumulation increased in the medium containing L-glutamic acid . Ornithine carbamoyltransferase of this strain was genetically derepressed . These results suggested that carbamoylphosphate might be deficient in vivo . With the intention to increase endogenous carbamoylphosphate, pyrimidine analogs inhibiting growth were selected and the mutants resistant to these compounds were derived from the AHr-5 mutant . Of the resistant mutants derived, the 6-azauracil-resistant mutant AAr-9 produced 28 mg of L-arginine per ml, which corresponded to more than twofold the amount produced by the parent strain . Derivation of an arginine-requiring mutant from the double-resistant mutant AAr-9 provides a new advantageous method for the production of L-citrulline . The increase in arginine and citrulline production is discussed.

Mol Gen Genet, 1977 Nov 29, 157(2), 175 - 82
Molecular cloning and in vitro transcription of Bacillus subtilis plasmid in Escherichia coli; Horinouchi S et al.; A composite plasmid (pAT2010) has been constructed in vitro from RSF2124 and Bacillus subtilis IFO3022 plasmid (pAT1060) by covalent joining of the two DNA molecules by means of Escherichia coli DNA ligase through the cohesive ends generated by restriction endonuclease RI (EcoRI) cleavage . The composite plasmid was selected by transformation of E . coliC600r-m- with the ligated mixture after enrichment for composite plasmid by preparative agarose gel electrophoresis, and plating of the transformants on a medium containing ampicillin and colicin E1 . Treatment of the composite plasmid with EcoRI yielded two fragments corresponding to the linear forms of the parental plasmids . The composite plasmids replicated as biologically functionally units in E . coli, and expressed genetic information carried by RSF2124 . In the presence of chloramphenicol, the composite plasmids continued to replicate and the copy number gradually increased . Such nature of replication in the presence of chloramphenicol is characteristic to RSF2124 derived from colicin E1 factor, and so it is suggested that the replicator of RSF2124 is functional in the composite plasmid . The composite plasmid was found to synthesize mRNA of B . subtilis plasmid in cell-free extracts of E . Coli, by hybridization of the mRNA to the original plasmid DNA of pAT1060.

Biochemistry, 1977 Nov 15, 16(23), 5009 - 15
Involvement of precursor-specific segments in the in vitro maturation of Bacillus subtilis precursor 5S ribosomal RNA; Meyhack B et al.; In vitro maturation of precursor 5S ribosomal RNA (p5A) from Bacillus subtilis effected by RNase M5 yields mature 5S RNA (m5, 116 nucleotides), and 3' precursor-specific segment (42 nucleotides), and a 5' precursor-specific segment (21 nucleotides) (Sogin, M.L., Pace, B., and Pace, N.R . (1977), J . Biol . Chem . 252, 1350) . Limited digestion of p5A with RNase T2 introduces a single scission at position 60 of the molecule; m5 is cleaved at the corresponding nucleotide residue . The complementary "halves" of the molecules could be isolated from denaturing polyacrylamide gels . The isolated fragments of p5A are not substrates for RNase M5, suggesting that some recognition elements can be utilized by RNase M5 only when presented in double-helical form . In exploring the involvement of the precursor-specific segments in the RNase M5-p5A interaction, substrate molecules lacking the 3' or 5' precursor-specific segment were constructed by reannealing complementary "halves" from p5A and m5 RNA . The artificial substrate lacking the 5'-terminal precursor segment was cleaved very much more slowly than the lacking t' segment; the 5' precursor-specific segment therefore contains one or more components recognized by RNase M5 during its interaction with the p5A substrate.

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