Naturwissenschaften, 1979 Apr, 66(4), 182 - 9 Gentic engineering for practical application; Klingmuller W; Genetic engineering has ushered in a new era in biology . Although many problems are still to be solved, there are examples that point to a possible later application for the benefit of mankind: Bacteria can be manipulated to degrade crude-oil spillages, to produce human insulin and to bind nitrogen from the air . If all the bacteria that are indigenous to agricultural soils could be made to bind nitrogen, an increase in soil fertility might well result. J Bacteriol, 1979 Apr, 138(1), 40 - 7 Modulation of gene expression by drugs affecting deoxyribonucleic acid gyrase; Sanzey B; Nalidixic acid (Nal), a drug which affects deoxyribonucleic acid gyrase activity, inhibits the expression of catabolite-sensitive genes: the three maltose operons, the lactose and galactose operons, and the tryptophanase gene . A correlation between the degree of sensitivity to Nal and that to catabolite repression has been observed . The expression of the threonine and tryptophan operons, insensitive to catabolite repression, is insensitive to Nal . The expression of the lacZ gene under the control of the IQ promoter is activated by Nal . Strains carrying a mutation in the nalA locus are resistant to these effects . Novobiocin, which inhibits the negative supercoiling activity of deoxyribonucleic acid gyrase, affects expression of the operons similarly to Nal . The involvement of promoters in Nal and novobiocin action, as well as a possible role of in vivo negative supercoiling in the selectivity of gene expression, are discussed. Nucleic Acids Res, 1979 Apr, 6(4), 1521 - 34 Pyrophosphate-condensing activity linked to nucleic acid synthesis; Volloch VZ et al.; In some preparations of DNA dependent RNA polymerase a new enzymatic activity has been found which catalyzes the condensation of two pyrophosphate molecules, liberated in the process of RNA synthesis, to one molecule of orthophosphate and one molecule of Mg (or Mn) - chelate complex with trimetaphosphate . This activity can also cooperate with DNA-polymerase, on condition that both enzymes originate from the same cells . These results point to two general conclusions . First, energy is conserved in the overall process of nucleic acid synthesis and turnover, so that the process does not require an energy influx from the cell's general resources . Second, the synthesis of nucleic acids is catalyzed by a complex enzyme system which contains at least two separate enzymes, one responsible for nucleic acid polymerization and the other for energy conservation via pyrophosphate condensation. Nucleic Acids Res, 1979 Apr, 6(4), 1309 - 21 A new method for the size estimation of the RNA genome segments of influenza virus; Sleigh MJ et al.; Previous estimates of the size of the RNA genome segments of influenza virus have been unreliable because of a lack of suitable RNA species as size markers . We have attempted to overcome this problem by utilising the ability of AMV reverse transcriptase to synthesise full length DNA copies of RNA molecules in the presence of a suitable primer . By comparing such DNA copies of the RNA segments of the influenza virus genome with sequenced restriction fragments from the E . coli plasmid pBR322, we have made more reliable estimates of the sizes of the eight genome segments from influenza virus A/NT/60/68. Nucleic Acids Res, 1979 Apr, 6(4), 1221 - 39 Polyadenylation and reverse transcription of influenza viral RNA; Emtage JS et al.; The polyadenylation of Fowl Plague Viral RNA and of Influenza A/Victoria Viral RNA using E . coli poly (A) polymerase and the subsequent reverse transcription of the polyadenylated species is reported . We have shown that all 8 genome fragments are adenylated and that an average of 25--30 adenylic acid residues per molecule is sufficient for maximal transcription with reverse transcriptase . The cDNA product is 95% sensitive to Sl-nuclease and hybridisation analysis against viral RNA reveals it to be a faithful copy of the RNA . Amongst the transcription products are long, discrete copies of genes 1--8, the lengths of which are comparable with those of the vRNA determined by electrophoresis on formamide acrylamide gels . These single-stranded cDNAs have been further transcribed to form double-stranded products with hair-pin structures at one end . Analysis of this material on native acrylamide gels revealed some DNA bands corresponding to the predicted sizes for genes 4--8. Biochim Biophys Acta, 1979 Mar 29, 572(3), 472 - 82 Glycerol 3-phosphate analogues as metabolic inhibitors in Escherichia coli, 3-hydroxy-4-oxobutyl-1-phosphonate, a drug that interferes with normal phosphoglyceride metabolism; Tang CT et al.; 3-Hydroxy-4-oxobutyl-1-phosphonate, the phoshonic acid analogue of glyceraldehyde 3-phosphate, enters Escherichia coli via the glycerol 3-phosphate transport system . There is no differential effect upon the accumulation of deoxyribonucleic acid, ribonucleic acid, or phosphoglycerides, although the accumulation of proteins was less effected . Examination of the phospholipids revealed that phosphatidylglycerol accumulation was most severely inhibited and cardiolipin accumulation was least affected . Concentrations of glyceraldehyde 3-phosphate and its phosphonic acid analogue that markedly inhibit macromolecular and phosphoglyceride biosynthesis have no effect upon the intracellular nucleoside triphosphate pool size . The phosphonate is a competitive inhibitor of sn-glycerol 3-phosphate in reactions catalyzed by acyl coenzyme A:sn-glycerol-3-phosphate acyltransferase and CDP-diacylglycerol:sn-glycerol-3-phosphate phosphatidyltransferase . A Km mutant for the former enzyme was susceptible to the phosphansferase activity . Studies with mutant strains ruled out the aerobic glycerol-3-phosphate dehydrogenase, glycerol-3-phosphate synthase, and fructose-1,6-biphosphate aldolase as the primary sites of action. Biochim Biophys Acta, 1979 Mar 28, 562(1), 51 - 61 DNA synthesis with methylated poly(dC-dG) templates . Evidence for a competitive nature to miscoding by O(6)-methylguanine; Abbott PJ et al.; The alternating copolymer poly(dC-dG) has been methylated with either dimethyl sulphate or N-methyl-N-nitrosourea and the levels of the various methylation products determined . In addition to the 3-methylcytosine, 3-methylguanine and 7-methylguanine (produced by both agents) reaction with N-methyl-N-nitrosourea also yielded easily detectable amounts of O(6)-methylguanine and phosphotriesters . These methylated polymers were then used as templates in an in vitro assay with Escherichia coli DNA polymerase I measuring the incorporation of complementary (dCMP and dGMP) and noncomplementary (dAMP and dTMP) nucleotides . When the dimethyl sulphate-methylated polymer was used as template there was virtually no detectable incorporation of non-complementary nucleotides indicating that no miscoding could be attributed to the presence of 3-methylcytosine, 3-methylguanine or 7-methylguanine . However, when the N-methyl-N-nitrosourea-methylated polymer was used as template there was a specific incorporation of dTMP but not of dAMP . The amount of dTMP incorporated was always less than the level of O(6)-methylguanine in the template and was found to vary with the relative concentrations of the deoxynucleoside 5'-triphosphates in the assay . As the amount of dCTP present in the assay was decreased the wrong incorporation of dTMP increased and approached the level that would have been expected for a one-to-one miscoding by O(6)-methylguanine as the concentration of dCTP approached zero . The results indicate that O(6)-methylguanine is capable of miscoding with a DNA polymerase but the miscoding is competitive with the normal incorporation of dCMP: when the 5'-triphosphate precursors are present in equal amounts approximately one O(6)-methylguanine in three miscodes leading to the incorporation of dTMP. Biochim Biophys Acta, 1979 Mar 28, 562(1), 112 - 30 Specificity of chromatin transcription in vitro . Anomalies due to RNA-dependent RNA synthesis; Pays E et al.; In the presence of Mn2+, globin mRNA can be transcribed into a partial RNA copy by Escherichia coli RNA polymerase . This process also occurs when the mRNA is transcribed together with chromatin . A fraction, at least, of the newly synthesized RNA copy (anti-globin RNA) can serve as a template for the synthesis of globin sequences of the same polarity as the original mRNA . This process is sufficient to explain the specific synthesis of a subset of the globin RNA on mouse foetal liver chromatin . It also accounts for the synthesis of double-stranded RNA sequence by E . coli RNA polymerase, on chromatin as well as on pure mRNA . Results are presented suggesting that the poly(A) tract of the mRNA could be preferentially transcribed . In the presence of Mg2+, the RNA-dependent transcription is strongly inhibited, as well as the synthesis of double-stranded RNA . Under these conditions, the transcription on chromatin appears to be largely DNA dependent, and the synthesis of globin sequences is completely asymmetric . Spermine (0.3 mM) seems to improve the specificity of transcription . The transcription of chromatin in vitro is thus largely dependent on the nature of the divalent cation present in the in the incubation mixture. Mol Gen Genet, 1979 Mar 27, 171(3), 261 - 75 Threonine deaminase: autogenous regulator of the ilv genes in Escherichia coli K-12; Abrescia P et al.; In this paper we analyze the effect of mutations in three genes, ilvO, ilvA and rho, on the expression of the ilvEJGDA gene cluster of Escherichia coli K-12 . The ilvO603 mutation causes a cis-dominant derepression of the ilvEJGDA genes . In particular, the ilvG gene, not expressed in the wild type, becomes expressed in the ilvO603 strain . We have introduced ilvA mutations (ilvA454 or ilvA628) in the ilvO603 strain and we show that ilvG expression requires the presence in cis of both an ilvO603 mutation and of an ilvA+ allele . The ilvG gene is not expressed when in trans is present an ilvO+, ilvA+ genotype . However, it is expressed when the chromosome in trans is ilvO603, ilvA+ (ilvG-) . We suggest that ilvO603 is part of ilvA, the structural gene for threonine deaminase, and that threonine deaminase from the ilvO603 mutant binds the ilvO603 site and not the ilvO+ site . Therefore, the ilvA gene product would be a cis-acting protein . Mutations in the rho gene cause derepression of the ilvEJGDA gene cluster without a concomitant expression of the ilvG gene . We show that introduction of either a rho-218 or a rho-115 mutation into the ilvO603, ilvA454 double mutant causes expression of ilvG . We therefore suggest that the ilvA gene product, threonine deaminase, is involved in termination of transcription as an antagonist of the rho gene product . Introduction of ilvA454 into an ilvO603 strain causes also a decrease in expression of the ilvE, ilvJ and ilvD genes . This effect is maximum in the case of the ilvD gene and we studied it in detail in isogenic strains containing also the rho-218 mutation. Mol Gen Genet, 1979 Mar 27, 171(3), 251 - 6 Pathways for repair of DNA damaged by alkylating agent in Escherichia coli; Yamamoto Y et al.; A strain with both the polA12 and the alk-1 mutation is only slightly more sensitive to methyl methane sulfonate (MMS) than isogenic strains with only one of the mutations . On the other hand, alk-1 recA1 double mutant is much more sensitive to MMS than are strains carrying either one of alk or recA mutation . It was suggested that the alk and the polA gene products are involved in the same DNA repair process whereas the recA function is independent from the process . The yield of MMS-induced mutation (Arg- (argE) to Arg+ reversion) in alk mutant is considerably higher than that in wild type strain . Thus, the repair process in which the alk gene product is involved is relatively accurate . When MMS-treated lambda phages were plated on MMS-treated bacteria, there were considerable increases in survival of treated phage even in recA alk double mutant . It seems that a new repair pathway, which is specific for alkylating agent-induced damages and is not dependent on the RecA function, may be induced on exposure of bacteria to the alkylating agent. J Biol Chem, 1979 Mar 25, 254(6), 1902 - 12 Error induction and correction by mutant and wild type T4 DNA polymerases . Kinetic error discrimination mechanisms; Clayton LK et al.; The fidelity of DNA synthesis as determined by the misincorporation of the base analogue 2-aminopurine in competition with adenine has been measured as a function of deoxynucleoside triphosphate substrate concentrations using purified mutator (L56), antimutator (L141), and wild type (T4D) T4 DNA polymerases . Although the rates of both incorporation and turnover of aminopurine and adenine decrease as substrate concentrations are decreased, the ratio of turnover/polymerase activity is increased . Thus, the nuclease/polymerase ratio of each of these three DNA polymerases can be controlled . The misincorporation of aminopurine decreases with decreasing substrate concentrations such that all three enzymes approach nearly identical misincorporation frequencies at the lowest substrate concentration . The increased accuracy of DNA synthesis corresponds to conditions producing a high nuclease/polymerase ratio . The misinsertion frequency for aminopurine is independent of substrate concentrations and enzyme phenotype; therefore, the increased accuracy of DNA synthesis with decreasing substrate concentrations is shown to be a result of increased nuclease activity and not increased polymerase or nuclease specificity . The data are analyzed in terms of a kinetic model of DNA polymerase accuracy which proposes that discrimination in nucleotide insertion and removal is based on the free energy difference between matched and mismatched base pairs . A value of 1.1 kcal/mol free energy difference, delta G, between adenine: thymine and aminopurine:thymine base pairs is predicted by model analysis of the cocentration dependence of aminopurine misincorporation and removal frequencies . An independent estimate of this free energy difference based on the 6-fold higher apparent Km of T4 DNA polymerase for aminopurine compared to adenine also gives a value of 1.1 kcal/mol . It is shown that the aminopurine misinsertion frequency for an enzyme having either extremely low 3'-exonuclease activity, Escherichia coli DNA polymerase I, or no measurable exonuclease activity, calf thymus DNA polymerase alpha, is 12 to 15%, which is similar to that for the T4 polymerases and consistent with delta G approximately 1.1 kcal/mol. J Biol Chem, 1979 Mar 25, 254(6), 1761 - 4 Methanol formation in vivo from methylated chemotaxis proteins in Escherichia coli; Toews ML et al.; Chemotactically wild type Escherichia coli were incubated with L-{methyl-3H}methionine to label the methyl groups of their methyl-accepting chemotaxis proteins . Cells were then treated to specifically demethylate these proteins . We have identified the end product of this demethylation as {3H}methanol in the cell-free medium from treated cells. J Biol Chem, 1979 Mar 25, 254(6), 1951 - 6 Identification of a novel RNA molecule in a new RNA processing mutant of Escherichia coli which contains 5 S rRNA sequences; Ghora BK et al.; A temperature-sensitive mutant of Escherichia coli at the nonpermissive temperature fails to produce normal levels of 5 S rRNA . Instead, a number of larger RNA molecules are accumulated . One of these molecules, a 9 S RNA, contains 5 S rRNA sequences . When the strain is shifted from a nonpermissive to a permissive temperature, radioactive label is lost from the 9 S RNA and appears in 5 S rRNA . The identification of this 5 S rRNA-containing molecule indicates the participation of a new processing ribonuclease (RNase E) in the maturation of rRNA in E . coli . The 9 S RNA was not detected in a wild type strain, indicating that the processing step(s) involved in the formation of 5 S rRNA might be performed before the growing rRNA transcript is terminated. J Biol Chem, 1979 Mar 25, 254(6), 1778 - 80 31P NMR of alkaline phosphatase . Saturation transfer and metal-phosphorus coupling; Otvos JD et al.; The rate constants which characterize the formation and breakdown of the noncovalent (E.P) and covalent (E-P) enzyme-phosphate intermediates on the alkaline phosphatase reaction pathway are known to be sensitive to the nature of the metal ion bound to the enzyme . 31P NMR saturation transfer has been demonstrated to provide a simple and sensitive method for measuring the metal ion dependence of these rates under equilibrium conditions . When the native Zn2+ was replaced by Cd2+, the 31P NMR spectrum at high pH revealed a new resonance at 12.6 ppm which has been assigned to the noncovalent enzyme.phosphate complex . Reconstituting the enzyme with enriched 113Cd2+ caused this unusually downfield-shifted resonance to appear as a doublet due to 113Cd-31P spin coupling (2J31P-O-113Cd = 30 Hz) . This result provides the first unequivocal evidence for direct metal-phosphate interaction in alkaline phosphatase. Biochim Biophys Acta, 1979 Mar 23, 552(1), 89 - 102 Small angle x-ray scattering from the inner and outer membranes from Escherichia coli; Harder ME et al.; Small angle X-ray data from purified forms of inner or cytoplasmic and outer membranes from Escherichia coli have been obtained and appear to be qualitatively similar . Transitory changes are apparent in the circularly averaged X-ray profiles from inner membranes . Such results could be due to the loss or denaturation of peripheral membrane proteins . Some partially dried forms of outer membrane are partly ordered and produce diffraction patterns which support an underlying bilayer structure . An extra light membrane fraction which results from membrane preparations utilizing a French pressure cell for spheroplast disruption has been characterized and shown to be similar to inner membrane . The purified membranes produce small angle X-ray diffraction patterns which are much different from those of lipid dispersions and the differences are attributable to the high protein content of the intact membranes . While the small angle X-ray region may be useful for characterizing the membrane preparations, the paucity of detail in the diffraction pattern suggest that it will be of little value in describing the complex underlying membrane structure. Mol Gen Genet, 1979 Mar 20, 171(2), 215 - 8 Study of a suppressor of mutT in Escherichia coli; Ray U; A suppressor mutation of E . coli mut T has been isolated and mapped in the lue--ace(E/F) region. Mol Gen Genet, 1979 Mar 20, 171(2), 193 - 203 Transposon-insertion mutants of Escherichia coli K12 defective in a component common to galactose and ribose chemotaxis; Harayama S et al.; From a collection of 8,000 transposon-insertion mutants of Escherichia coli K12 we identified two mutations, trg-1::Tn5 and trg-2::Tn10, that simultaneously eliminate chemotactic response to ribose and galactose, two attractants recognized by independent receptors . We show that these transposon-insertions confer a Trg phenotype, indicating that this specific pattern of tactic defects is a null phenotype . The two mutation sites are cotransductionally linked to an extend consistent with placement in the same gene . The Trg phenotype of a family of deletion mutants produced by curing trg-2::Tn10 implies that trg is a single gene . Experiments with appropriate F-primes and Hfr's locate the trg locus at approximately 31 min on the linkage map, with a marker order: pyrF-rac-(P.O . 43)-trg-man . We also found one trg mutant whose Trg phenotype was not linked to a transposon-insertion but is probably the result of a mutator activity in the parent strain . Selection of transposon-insertions near, but not in trg allowed demonstration of a very close linkage between the spontaneous trg-3 and the transposon-generated trg's, indicating all three mutations are probably in the same gene . In our manipulations of transposon-insertions we found that Tn5 had a tendency to translocate from its initial site of insertion while Tn10 was relatively stable . The trg-product is probably a chemotactic signal transducer, which interacts directly with two independent receptor proteins and transmits information to the central chemotactic machinery. Mol Gen Genet, 1979 Mar 20, 171(2), 135 - 43 Genetic evidence for the nature, and excision repair, of DNA lesions resulting from incorporation of 5-bromouracil; Krych M et al.; Escherichia coli mutants defective in DNA uracil N-glycosidase (ung-) or endonuclease VI active against apurinic/apyrimidinic sites in DNA (xthA-) exhibit enhanced sensitivity towards 5-bromodeoxyuridine relative to the wild type strain, pointing to involvement of these enzymes in repair of bromouracil-induced lesions in DNA . Mutants defective in DNA polymerase I, either in polymerizing activity (polAl-) or (5' leads to 3')-exonuclease activity (polA107-) exhibit unusually high sensitivity (including marked lethality) in the presence of 5-bromodeoxyuridine . The results indicate that DNA polymerase I, and its associated (5'--3')-exonuclease activity, are involved in repair of bromouracil-induced lesions and are not readily replaced, if at all, by DNA polymerases II and III . Thermosensitive mutant in DNA ligase gene (lig ts7) shows high sensitivity towards 5-bromodeoxyuridine at 42 degrees C indicating the role of the enzyme in repair of bromouracil-induced lesions in DNA . Involvement of DNA uracil N-glycosidase, and endonuclease active against apurinic/apyrimidinic sites in recognition and repair of 5-bromouracil-induced damage permits of some inferences regarding the nature of this damage (lesions), in particular dehalogenation of incorporated bromouracil to uracil residues. Biochemistry, 1979 Mar 20, 18(6), 979 - 83 Subunits of RNA polymerase in function and structure . 8 . Catalytic properties of self-reactivated core enzyme; Saitoh T et al.; As an attempt to identify the maturation pathway of Escherichia coli RNA polymerase, the catalytic properties of core enzyme reactivated in the absence of maturation-promoting factors (sigma subunit or DNA) (that is, of self-reactivated core enzyme) were compared with those of native core enzyme . Differences have been found in the intrinsic activities such as in the template specificity, Km value of DNA template for the polymerase, activation energy for RNA synthesis, and increment of enzyme activity by sigma subunit . These observations imply that the transcription initiation by self-reactivated core enzyme is inaccurate and, therefore, more strict conditions including the presence of maturation-promoting factors are required for premature core be activated to the genuine function with the transcription specificity of native core enzyme. Biochemistry, 1979 Mar 20, 18(6), 972 - 8 Subunits of RNA polymerase in function and structure . 7 . Structure of premature core enzyme; Ishihama A et al.; The structure of premature core enzyme, an obligatory intermediate in both in vivo and in vitro assembly of Escherichia coli DNA-dependent RNA polymerase, was compared with that of native core enzyme . Though this assembled but inactive form of core enzyme harbors the gross conformation similar to that of native enzyme, minor and presumably local differences exist, which were identified by near-ultraviolet circular dichroism spectra, tritium-hydrogen exchange rate, protease sensitivity, intersubunit cross-linking rate by bifunctional reagents, sedimentation behavior, and elution profile from phosphocellulose . Taken together these results indicate that the core enzyme subunits are loosely associated in the premature core . The temperature-dependent maturation is required for the core subunits to be tightly associated, leading to the formation of structurally stable and functionally active RNA polymerase. Biochemistry, 1979 Mar 20, 18(6), 1063 - 7 Lac UV5 transcription in vitro . Rate limitation subsequent to formation of an RNA polymerase-DNA complex; Stefano JE et al.; The kinetics of transcription of lac UV5 mRNA using purified DNA restriction fragment as template has been studied . This template, which contains only 203 base pairs, directs the formation of a 67-base lac mRNA with high specificity . The half-time for formation of a DNA-RNA polymerase complex is approximately 0.2 min . However, upon addition of 200 micron nucleoside triphosphates to this complex, RNA production proceeds with a half-time of approximately 1 min . Therefore, it is suggested that the rate-limiting step for lac UV5 mRNA production, under typical in vitro conditions, occurs subsequent to the formation of a promoter-specific complex. Eur J Biochem, 1979 Mar 15, 95(1), 39 - 49 Degradation of the DNA-binding domain of wild-type and i-d lac repressors in Escherichia coli; Schlotmann M et al.; It has been shown that 28 transdominant mutant lac repressors which have lost operator DNA-binding ability in vivo and in vitro, but still bind inducer and are able to form tetramers (i-d repressors), could be divided into two groups by their capacity or incapacity to bind non-specifically to the phosphate groups of the DNA backbone . All but one of 15 analysed i-d repressors with amino acid substitutions to the C-terminal of residue 52 showed uneffected non-specific DNA binding . All 13 tested i-d repressors with amino acid substitutions to the N-terminal of residue 53 did not bind to double-stranded DNA, and 11 of these repressors derived from missense mutations in the lacI gene were endogenously degraded . The degradation in vivo only affects the amino-terminal 50-60 residues producing a mutant-specific pattern of stable repressor fragments . These fragments are tetrameric and capable of binding inducer in vivo and in vitro . The proteolytic attack presumably takes place during synthesis of the i-d repressors, since the resulting fragments are stable, both in vivo (as shown by a pulse-chase experiment) and in vitro . The proteolysis in vivo depends on the growth conditions of the bacteria and is higher in cells grown in minimal media than in rich media . Wild-type repressor is only susceptible to limited proteolysis in cells grown in minimal media but not in cells grown in rich media . The results suggest that the majority of the sequence alterations before residue 53 in missense mutant i-d lac repressor proteins affect the three-dimensional structure of the amino-terminal DNA-binding domain of the repressor protein, making it susceptible to proteolytic attack by one or several intracellular proteases. J Mol Evol, 1979 Mar 15, 12(3), 189 - 95 A unique pattern of toxic synthesis in pentitol catabolism: implications for evolution; Scangos GA et al.; All of our Escherichia coli C mutants blocked in the first step of D-arabitol catabolism (D-arabitol dehydrogenase) became unable to grow in the presence of D-arabitol . We have shown that this sensitivity is eliminated by a defect in the second enzyme of the pathway (D-xylulokinase), leading to a pattern of toxicity and its relief which has not been previously reported . We have found a similar pattern of toxicity and its relief in the closely related ribitol pathway . The evolutionary significance of these findings is discussed. Int J Cancer, 1979 Mar 15, 23(3), 344 - 52 Characteristics of a low-molecular-weight factor extracted from mouse tumors that affects in vitro properties of macrophages; Cheung HT et al.; A dialysable low-molecular-weight factor capable of affecting in vitro properties of macrophages was extracted from four different mouse tumors . This factor not only modulates closely related properties of peritoneal macrophages such as spreading and migration but also inhibits lipopolysaccharde-induced tumoricidal activity of these cells . It can be extracted not only from tumor tissues but also from tumor cells grown in vitro . The appearance of this factor is unique to tumors and it is not present in detectable quantities in normal tissues . The factor from one of the tumors, Lewis lung carcinoma, was purified extensively and the partially purified factor retains all the above effects on macrophages . It is not sensitive to pronase or a mixture of bovine spleen phosphodiesterase II, E . coli alkaline phosphatase and pancreatic ribonuclease . The factor is lipid-like in character and it is soluble in both organic solvents and aqueous media . It has ionizable group(s) and is anionic at neutral pH but non-ionic under acidic conditions. Chromosoma, 1979 Mar 12, 71(3), 263 - 81 The organization of the ribosomal RNA genes of Chironomus tentans and some closely related species; Degelmann A et al.; Southern gel analysis of total DNA from Chironomus tentans showed that the rRNA genes (rDNA) are homogeneous in structure . After cloning in Escherichia coli plasmid pBR313, the rDNA organisation was further studied by restriction fragment analysis and R-loop mapping . No heterogeneity could be detected by heteroduplex analysis of six different cloned rRNA cistrons . R-loop sizes of 1.69 and 3.63 kilobases (kb) were measured for the 18S and 28S rRNA coding sequences . The two spacers are 0.75 and 1.77 kb long . Southern gel analysis showed also a homogeneous rDNA structure for a Canadian population of C . tentans and C . pallidivittatus . The same technique indicated, however, that the rDNA of two other closely related species of C . thummi and C . melanotus is heterogeneous in structure . A possible correlation between this heterogeneity and the presence of heterochromatin in these species is discussed. Med J Aust, 1979 Mar 10, 1(5), 154 - 5 Coliform status of domestic swimming pools; Gluer J et al.; An investigation of maintenance, usage, and coliform status of water in 104 domestic swimming pools in 13 Brisbane suburbs was made in January, 1978 . The results showed that domestic pools are often imperfectly serviced, and are potential infection sources . The need for health education of pool owners, and for more reliable methods of pool sanitization is emphasized. J Biol Chem, 1979 Mar 10, 254(5), 1748 - 53 DNA polymerase III of Escherichia coli . Purification and identification of subunits; McHenry CS et al.; DNA polymerase III, the core of the DNA polymerase III holoenzyme, has been purified 28,000-fold to 97% homogeneity from Escherichia coli HMS-83 . The enzyme contains subunits: alpha, epsilon, and theta of 140,000, 25,000, and 10,000 daltons, respectively . The alpha subunit has been previously shown to be a component of both DNA polymerase III and the more complex DNA polymerase III holoenzyme (Livingston, D.M., Hinkle, D., and Richardson, C . (1975) J . Biol . Chem . 250, 461-469; McHenry, C., and Kornberg, A . (1977) J . Biol . Chem . 252, 6478-6484) . It is demonstrated here that the epsilon and theta subunits are also subunits of the DNA polymerase III holoenzyme . Thus, the DNA polymerase III holoenzyme contains at least six different subunits . Our preparation has both the 3' leads to 5' and 5' leads to 3' exonuclease activities previously assigned to DNA polymerase III (Livingston, D., and Richardson, C . (1975) J . Biol . Chem . 250, 470-478). J Biol Chem, 1979 Mar 10, 254(5), 1699 - 706 The structure of cytochrome b562 from Escherichia coli at 2.5 A resolution; Mathews FS et al.; The structure of cytochrome b562 from Escherichia coli has been determined at 2.5 A resolution by x-ray diffraction methods . Protein phases were computed by the single isomorphous replacement method with anomalous scattering measurements from the native and uranyl acetate-substituted crystals . The electron density was averaged about the noncrystallographic 2-fold axis relating 2 molecules in the triclinic unit cell . The protein consists of four nearly parallel alpha helices and represents a new class of cytochrome structure . The heme group is inserted between the helices near one end of the molecule with one heme face partially exposed to solvent . The two heme ligands are histidine and methionine . The 2 phenylalanines are packed internally near the heme group, and the 2 tyrosines are on the surface, also near the heme group . The folding of the protein resembles that of hemerythrin and tobacco mosaic virus protein and shows a different topology from that of cytochrome b5, cytochrome c, or the globins. J Biol Chem, 1979 Mar 10, 254(5), 1457 - 61 Subunit structure of functional porin oligomers that form permeability channels in the other membrane of Escherichia coli; Nakae T et al.; Oligomers of a protein, porin, form permeability channels in the outer membrane of Escherichia coli B . A functional porin oligomer was identified and was purified to homogeneity by gel filtration in the presence of salts and sodium dodecyl sulfate . Molecular weights of purified porin oligomer and heat-dissociated monomer appeared to be 102,900 and 32,600, respectively, when determined by sedimentation equilibrium in the presence of sodium dodecyl sulfate . We concluded that the porin oligomer thus consists of three identical subunits . These data and results from other laboratories suggest porin trimers exist also in the outer membrane of intact cells, and participate in the formation of permeability channels . It was found that porin trimer bound less sodium dodecyl sulfate than the porin monomer. Mol Gen Genet, 1979 Mar 9, 171(1), 75 - 8 Molecular cloning of the 4.2 Md EcoRI fragment of Aspergillus nidulans mitochondrial DNA; Bartnik E et al.; The 4.2 Md EcoRI fragment of Aspergillus nidulans mitochondrial DNA was cloned using the plasmid pBR 332 as vector and E . coli as host . Hitherto unknown sequence of HindIII sites within this region of mitochondrial genome was established. Mol Gen Genet, 1979 Mar 9, 171(1), 7 - 13 Transposition of DNA inserted into deletions of the Tn5 kanamycin resistance element; Meyer R et al.; Tn5-trp hybrid transposons have been constructed by insertion of a trpPOED Hind III fragment into an in vivo Tn5 internal deletion mutant or by substitution of trp for the internal Tn5 Hind III fragment . These hybrids are called, respectively, Tn409 and Tn410 . Both Tn409 and Tn410 will transpose into lambda in the presence of a complementing Tn5 element . In the absence of a wild Tn5, lysogens carrying R1162::Tn409 and R1162::Tn410 plasmids will yield lambdatrp phages at less than six per cent of the complemented frequency . This reduction indicates that Tn409 and Tn410 lack a diffusible transposition function provided by wild Tn5 elements . However, the formation of lambdatrp phages without complementation is real . Most of these transducing particles contain Tn409 and Tn410 still linked to the carrier R1162 plasmid . This observation suggests that uncomplemented Tn409 and Tn410 elements mediate the formation of lambda-transposon-plasmid cointegrate structures . Thus, the missing transposition function may be involved in resolving these cointegrate structures to the final lambda::Tn409 or lambda::Tn410 product. Mol Gen Genet, 1979 Mar 9, 171(1), 53 - 8 DNA synthesis in UV-irradiated Escherichia coli K-12 strains carrying dnaA mutations; Jonczyk P et al.; It has been found that UV irradiation of dnaA mutants of E . coli K-12 enables the initiation of DNA synthesis at a temperature restrictive to these mutants . The UV-induced DNA synthesis is dependent on protein synthesis and on a transcriptional event at a time when protein synthesis is no longer required . In contrast to dnaA mutants UV irradiation fails to induce DNA synthesis in the two other initiation mutants dnaC2 and dnaB252 . DNA synthesis at the restrictive temperature is initiated also when the tif-1 phenotype is expressed in the dnaA46 tif-1 double mutant . Possible mechanisms of the observed capability of dnaA mutants to synthesize DNA at the restrictive temperature after UV irradiation or under conditions of tif-1 expression are discussed. Mol Gen Genet, 1979 Mar 9, 171(1), 1 - 6 Spontaneous degradation of pRD1 DNA into unique size classes is recA dependent; Puhler A et al.; The his and nif genes of the P1 type plasmid pRD1 were lost at a high frequency in a recA+ but not in a recA- Escherichia coli host during growth in a non-selective medium . 92% of the His- Nif- segregants after 6 subcultures retained the genetic markers of the precursor plasmid RP4, while the remainder lost all of the pRD1 markers with the concomitant loss of ccc-DNA . Plasmids purified from the His- Nif- segregants resembled RP4 in the physical and genetic properties examined . The contour length of pRD1 DNA molecules from a recA- strain was 49 micrometer corresponding to a molecular weight of 101 Mdals and the buoyant density was 1.715 g/cm3 . In contrast, the contour lengths of plasmid molecules isolated from a recA+ host carrying pRD1 fell into 3 size classes of 49, 19 and 2 micrometer corresponding to molecular weights of 101, 39 and 4 Mdals respectively and two DNA species of buoyant density 1.715 and 1.719 g/cm3 were observed. Science, 1979 Mar 9, 203(4384), 1014 - 6 Binding of cis- and trans-dichlorodiammineplatinum(II) to DNA: evidence for unwinding and shortening of the double helix; Cohen GL et al.; The antitumor drug cis-dichlorodiammineplatinum(II) (cis-DDP) and the inactive trans isomer bind and produce cooperative changes in closed and nicked circular duplex DNA's . Covalent binding of both platinum complexes to the closed circular DNA alters the degree of supercoiling, presumably by disrupting and unwinding the double helix . Electron micrographs show the platinated DNA's to be shortened by up to 50 percent of their original length . At similar ratios of bound platinum per nucleotide, the electrophoretic mobilities of the DNA's in gels containing the dye ethidium bromide are the same for both isomers . The only detectable difference in the binding of the two platinum isomers is an increase in the electrophoretic mobility in nondye gels of closed circular DNA having small amounts of bound cis-DDP that is not apparent for the trans complex. Mol Gen Genet, 1979 Mar 9, 171(1), 15 - 22 A missense mutation in the gene coding for ribosomal protein S17 (rpsQ) leading to ribosomal assembly defectivity in Escherichia coli; Herzog A et al.; The conditionally lethal mutation, 2861 mis, has been mapped inside the ribosomal protein gene cluster at 72 minutes on the Escherichia coli chromosome and was found to cotransduce at 97% with rpsE (S5) . The 2861 mis mutation leads to thermosensitivity and impaired assembly in vivo of 30S ribosomal particles at 42 degrees C . The strain carrying the mutation has an altered S 17 ribosomal protein; the mutational alteration involves a replacement of serine by phenylalanine in protein S 17 . Spontaneous reversion to temperature independence can restore the normal assembly in vivo of 30 S ribosomal subunits at 42 degrees C and the normal chromatographical behaviour of the S 17 ribosomal protein in vitro . We conclude therefore that the 2861 mis mutation affects the structural gene for protein S 17 (rpsQ). Science, 1979 Mar 9, 203(4384), 1012 - 4 beta-Galactosidase and selective neutrality; Holmquist R; Three hypotheses to explain the amino acid composition of proteins are inconsistent (P congruent to 10(-9) with the experimental data for beta-galactosidase from Escherichia coli . The exceptional length of this protein, 1021 residues, permits rigorous tests of these hypotheses without complication from statistical artifacts . Either this protein is not at compositional equilibrium, which is unlikely from knowledge about other proteins, or the evolution of this protein and its coding gene have not been selectively neutral . However, the composition of approximately 60 percent of the molecule is consistent with either a selectively neutral or nonneutral evolutionary process. MMW Munch Med Wochenschr, 1979 Mar 9, 121(10), 345 - 8 {Pathogenesis of coli enteritis (author's transl)}; Guggenbichler JP; The pathogenic mechanisms of E . coli in diarrheal diseases were largely obscure up to now . The discovery of an enterotoxin which corresponded with the cholera enterotoxin in its mode of action and is also largely identical with this molecule immunologically provided essentially new aspects of the pathogenic importance of E . coli in diarrheal diseases . The detection of enterotoxin formation depends on the biological test model and presently it is still expensive and unsuitable for routine laboratory use . The serological identification of coli bacteria is meaningless for the detection of enterotoxin formation . Enterotoxic coli are more frequently found (12%) in the large group of hemolytic organisms . In 1978 we isolated enterotoxic coli from the stools of 12 infants under one year old with stubborn diarrhea. Biochim Biophys Acta, 1979 Mar 8, 551(2), 238 - 47 Ionic selectivity of pores formed by the matrix protein (porin) of Escherichia coli; Benz R et al.; Incorporation of the matrix protein (porin) from the outer membrane of Escherichia coli into black lipid films results in the formation of ion-permeable pores with a single-pore conductance of the order of 2 nS (in 1 M KCl) . Information on the structure of this pore has been obtained by determining the selectivity for various species differing in charge and size . From the permeability of the pore for large organic ions (Tris+, glucosamine+, Hepes-) a minimum pore diameter of 0.8 nm is estimated . At neutral pH the pore is two to four times more permeable for alkali ions than for chloride . On the basis of the observed pH dependence of permeability, this cationic selectivity is explained by the assumption that the pore contains fixed negative charges. Mol Gen Genet, 1979 Mar 5, 170(3), 319 - 25 Studies of a plasmid coding for tetracycline resistance and hydrogen sulfide production incompatible with the prophage P1; Briaux S et al.; The plasmid pIP231, determining tetracycline resistance and hydrogen sulfide production is shown to belong to incompatibility group Y and to code for a restriction and modification system . Unlike the IncY plasmids, P7 and P15B, plasmid pIP231 shows only little genetic and physical homology with P1 prophage. Science, 1979 Mar 2, 203(4383), 887 - 92 Molecular cloning of polyoma virus DNA in Escherichia coli: lambda phage vector system; Chan HW et al.; The biological activity of recombinant phage and recombinant phage DNA containing monomeric or dimeric polyoma DNA inserts was examined in mice and cultured mouse cells . Recombinant preparations containing a single copy of viral DNA were invariably noninfectious; molecules containing a dimeric polyoma DNA insert were at least seven orders of magnitude less infectious than polyoma virions after parenteral inoculation . No infection was detected with any recombinant preparation after oral administration. Science, 1979 Mar 2, 203(4383), 883 - 7 Molecular cloning of polyoma virus DNA in Escherichia coli: plasmid vector system; Israel MA et al.; A series of recombinant plasmids containing polyoma virus (PY) DNA were constructed, and their biological activity was evaluated in mice and in cultured mouse cells . While all of the recombinants studied contain the complete, potentially infectious viral DNA, in no case was the intact recombinant PY-plasmid DNA, or live Escherichia coli containing the recombinant plasmids, capable of inducing PY infection of mice, either by feeding or by parenteral injection. Cell, 1979 Mar, 16(3), 535 - 49 Vitellogenin in Xenopus laevis is encoded in a small family of genes; Wahli W et al.; Vitellogenin, the yolk protein precursor, is produced in X . laevis liver from a 6.3 kilobase (kb) mRNA . Sequences of this mRNA have been transcribed into cDNA and cloned in E . coli . Some properties of 21 of these cloned DNAs, ranging in size from 1 to 3.7 kb, have been reported by Wahli et al . (1978b) . This paper reports restriction endonuclease mapping, cross hybridization, heteroduplex mapping in the electron microscope and heteroduplex melting experiments with these DNAs . We conclude that the cloned DNAs fall into two main groups of sequences which differ from each other in approximately 20% of their nucleotides . Each main group contains two subgroups which differ from each other by about 5% sequence divergence . By hybridizing cloned DNAs with restricted genomic DNA, we showed that sequences corresponding to all four sequence groups are present in a single animal . Furthermore, we have obtained tentative evidence for the presence of large intervening sequences in genomic vitellogenin DNA . Analysis of R loop molecules demonstrated that all four sequences are present in the vitellogenin mRNA population purified from individual animals . While some alternate explanations are not entirely excluded, we suggest that vitellogenin is encoded by a small family of related genes in Xenopus. J Lab Clin Med, 1979 Mar, 93(3), 437 - 48 Platelet and fibrinogen production: relative sensitivities to endotoxin; Alving BM et al.; The relative sensitivities of platelet and fibrinogen production to endotoxin were determined in male New Zealand rabbits . E . coli endotoxin was administered in single intravenous doses of 0.1 to 50.0 mu/g/kg body mass . 75SeM was injected 5, 12, or 18 hr after endotoxin, and the percent incorporation into platelets and fibrinogen was used to measure thrombopoiesis and fibrinogen synthesis . Leukopenia occurred after the infusion of endotoxin at all dose levels; the lowest dose that caused thrombocytopenia was 0.5 microgram/kg . Endotoxin was detected with the Limulus test in plasma of animals that received 5 to 50 microgram/kg . The stimulation of fibrinogen and platelet production, as well as the postinfusion decrease in platelet count, correlated directly with the log of the dose of endotoxin infused . The lowest dose of endotoxin that stimulated platelet and fibrinogen production was 0.5 microgram/kg . In animals that received this dose of endotoxin, increased fibrinogen production was detected only when 75SeM was injected 5 hr later . In animals injected 5, 12, or 18 hr later . Increased platelet production was detected at these two doses only when 75SeM was injected 18 hr after endotoxin . The data indicate that fibrinogen and platelet production have similar sensitivities to endotoxin, although the time course of stimulation is different for these two blood components. Immunopharmacology, 1979 Mar, 1(2), 125 - 35 Role of complement in endotoxin initiated lethality in mice; Curry BJ et al.; We have examined the role of complement in eliciting a lethal response in C3H/HeJ and C3H/St mice . The results reported here indicate that endotoxin-initiated complement activation, leading to significant drops in circulating C3 levels, is not sufficient to cause lethality . The complement system in both strains was demonstrated to be responsive in vitro to activation both by E . coli 0111:B4 LPS I, an alternative pathway activator in other systems, as well as S . minnesota R595 LPS, which activates almost exclusively the classical pathway . In vivo injection of high (lethal) doses of E . coli 0111:B4 LPS I and S . minnesota R595 LPS causes a significant decrease in the circulating C3 levels of both strains after 4 hr . In contrast, circulating C3 levels were not significantly different from normal values in either strain following injection with minimal (lethal) amounts of E . coli 0111:B4 LPS II, a weakly anticomplementary LPS preparation . In all cases, lethality was observed in only the C3H/St mice, indicating that neither complement activation, nor the lack of it, is responsible for lethality in mice. Neurosci Lett, 1979 Mar, 11(3), 279 - 82 Hypothalamic monoamine contents in endotoxin fever of new-born guinea pigs and kittens; Hahn Z et al.; In new-born guinea pigs and kittens Escherichia coli endotoxin did not significantly modify the hypothalamic level of either norepinephrine (NE) or 5-hydroxytryptamine (5-HT) . The hypothalamic 5-hydroxyindoleacetic acid (5-HIAA) content was significantly elevated in endotoxin-treated animals, but this elevation was confined to a period following the endotoxin injections by about 60 min in guinea pigs and 75-90 min in kittens. Ann Sclavo, 1979 Mar-Apr, 21(2), 235 - 41 {Comparative study of the different media of enrichment and isolation for the identification of enteric pathogens (author's transl)}; Mauro A et al.; In the present study, were compared several media in use in diagnostic laboratory for the isolation of enteric pathogens . The AA . have remarked the efficiency of Bacto-Selenite F and of Bacto-TT Broth Base Hajna, for the enrichment; it is suggested the use of Bacto Agar and of Bacto-Hektoen Enteric Agar for the isolation. Can J Genet Cytol, 1979 Mar, 21(1), 95 - 100 Reduction of sensitivity to ultraviolet light in excision-deficient Escherichia coli strain WP2 uvrA; Nestmann ER; Mutants with increased resistance to UV light were isolated from Escherichia coli WP2 uvrA . Most of these isolates were designated as true or partial revertants of the uvrA- mutation . However, one class of mutants showed only a moderate increase in resistance to UV . The gene responsible for this phenotypic change is closely linked to the lac locus and has been designated the suuA gene (suppressor of uvrA). Pediatr Res, 1979 Mar, 13(3), 148 - 51 Characteristics of impaired chemotactic function in cord blood leukocytes; Tono-Oka T et al.; Mobilities of cord blood granulocytes were studied using the agarose plate method and Boyden's chamber method . In the agarose plate, granulocytes of cord blood were shown to have moderately decreased responses in chemotaxis and chemokinesis induced by Escherichia coli-derived chemotactic factor and/or zymosan-activated serum, whereas they were shown to have a normal capacity of random mobility . Although their distance and index of chemotaxis or chemokinesis in the agarose plate were significantly less than those of adult granulocytes, response rate in both types of mobility were evidently higher compared with those in patients with chemotactic defect . Furthermore, there is a difference between chemotactic responses of cord blood granulocytes to E . coli-derived chemotactic factor and to zymosan-activated serum in the agarose plate method . Using the latter, a more distinguishable difference between chemotactic responses of cord blood granulocytes and adult granulocytes was shown . The Boyden's chamber method tended to show a more significant difference between chemotactic responses of granulocytes of cord blood and adults than in the agarose plate method. Gene, 1979 Mar, 5(3), 233 - 53 High frequency of yeast transformation by plasmids carrying part or entire 2-micron yeast plasmid; Gerbaud C et al.; By using two chimeric plasmids containing yeast ura3 gene and 2-micron yeast DNA linked to the bacterial plasmid pCR1, yeast transformation of a high frequency has been achieved . The first plasmid is such that the 2-micron DNA part, in which the ura3 gene is incorporated, can be removed in one step and thus the 2-micron-ura3 sequence can be considered as a "transposable" block . In contrast, the second one bears the entire 2-micron plasmid and the ura3 gene is inserted in the bacterial plasmid part . As shown through hybridization experiments and genetic studies, the ura3 gene was maintained as a cytoplasmic element . Plasmids recovered from the yeast transformants were used to transform Escherichia coli . Their analysis by EcoRI showed that in many cases the vector had recombined with the endogenous 2-micron DNA of the recipient strain . The specific activity of orotidine 5'-monophosphate decarboxylase (coded by ura3) in yeast transformants was 10- to 30-fold higher than in the wild type. Gene, 1979 Mar, 5(3), 207 - 31 Cloning and endonuclease restriction analysis of argF and of the control region of the argECBH bipolar operon in Escherichia coli; Crabeel M et al.; A 1.8 kb DNA fragment, liberated by endonuclease HindIII, contains the control region of the argECBH bipolar operon near one end and the weak secondary promoter of argH at the other extremity; it has been cloned in plasmid pBR322 . The same plasmid vector has been used to clone the argF gene liberated from the chromosome by endonuclease BamHI . Restriction patterns for the two hybrid plasmids have been determined, using enzymes AluI, BglI, EcoRI, HaeIII, HincII, HindIII, HpaI and II, PstI and SalI . Two AluI sites situated on either side of and close to a HincII target delineate two short fragments covering the whole of the argECBH control region . The argF control elements are located in a region accessible to further dissection by BamHI, EcoRI, PstI and HindIII . Carriers of the argF plasmid produce extremely high amounts of ornithine carbamoyltransferase, a feature useful for purification of this enzyme. J Gen Microbiol, 1979 Mar, 111(1), 193 - 200 Mutants of Escherichia coli K12 accumulating porphobilinogen: a new locus, hemC; McConville ML et al.; Mutants of Escherichia coli K12 which accumulated the haem precursor porphobilinogen are described . The mutants grew very slowly on carbon and energy sources which K12 uses only oxidatively, and they had low catalase activities, suggesting that they were deficient in haem . Extracts had one-tenth of the parental activity of the enzyme porphobilinogen deaminase . In transduction, the mutation mapped close to genes ilvD and metE at minute 84 . The gene was tentatively identified as hemC, coding for porphobilinogen deaminase . The gene symbol hemC replaces the earlier and temporary symbol popE. Infect Immun, 1979 Mar, 23(3), 690 - 9 Hemagglutination and adhesiveness of toxigenic Escherichia coli isolated from humans; Thorne GM et al.; Toxigenic strains of Escherichia coli isolated from humans were studied for adherence to human buccal mucosal epithelial cells . The E . coli strains were labeled with 3H-amino acids or fluorescein isothiocyanate . Toxigenic E . coli strains varied in their ability to adhere in the presence of mannose . Of 32 toxigenic strains examined, 52% bound to the buccal cells, whereas none of 8 control strains did so (Mann-Whitney U test, P =0.007) . The control strains were nontoxigenic E . coli isolates from humans, enterotoxigenic E . coli isolates from animals, and E . coli K-12 containing the K88 or K99 plasmid; these strains exhibited only background-level adherence in this assay . Among the toxigenic E . coli strains that bound to human buccal mucosal cells, there was no correlation with mannose-resistant hemagglutination (MR-HA) of guinea pig and human erythrocytes . Screening 32 strains, we found the following phenotypes: (i) MR-HA+, buccal adherent; (ii) MR-HA+, buccal nonadherent; (iii) MR-HA-, buccal adherent . Presumably the third group represents strains with another type(s) of surface attachment components not involved in the MR-HA reaction . Our findings indicate that a number of bacterial surface structures can function in MR-HA and buccal adherence. Appl Environ Microbiol, 1979 Mar, 37(3), 369 - 72 Induction of mutation in Escherichia coli by freeze-drying; Tanaka Y et al.; The effect of freeze-drying on phenotypic reversion of amino acid auxotrophy to prototrophy was studied in Escherichia coli . In a radioresistant strain, E . coli H/r 30 (uvr+ exr+), which can repair the deoxyribonucleic acid damaged due to freeze-drying, an increased mutation frequency from auxotrophy to prototrophy was observed with increased time of freeze-drying of the cells . On the other hand, in a radiosensitive strain, E . coli NG 30 (recA), which cannot repair the damaged deoxyribonucleic acid due to a lack of repair enzyme system, no significant reversion occurred, although the survival rate was very low . The rate of phenotypic reversion dut to freeze-drying in both E . coli RIMD 0509109 (uvr+ exr+) and RIMD 0509115 (uvr exr+) was almost the same, indicating that the phenomenon is independent of the uvr character . From these results it is concluded that mutation was induced in E . coli cells during the rehydration when the damaged deoxyribonucleic acid was repaired by exr character of the cells . Thus, we propose that a serious consideration should be paid to the freeze-drying technique to preserve bacterial cells. Lymphology, 1979 Mar, 12(1), 20 - 2 Qualitative and quantitative changes in thoracic duct lymph during canine experimental shock; Hayashi S et al.; Changes of the body fluid exchange and of the composition of metabolites in the hepatosplanchnic area in canine hemorrhagic and endotoxin or septic shock models were studied by investigating the qualitative and quantitative changes in thoracic duct lymph draining from abdominal organs . In the present study, it might be summarized that the changes in the flow rate and composition of thoracic duct lymph were put forward to much more directly and apparently indicate the degree of hepatosplanchnic cellular impairment in canine experimental shock than in the circulating blood. J Virol, 1979 Mar, 29(3), 1229 - 31 Development of Escherichia coli virus T1: escape from host restriction; Wagner EF et al.; The bacterial virus T1 grows interchangeably on different Escherichia coli strains (C, B, and K) . This implies that T1 has an efficient mechanism to overcome the host restriction barrier . The DNA of T1 was found to be methylated independently of the hosts . The percentage of N6-methyladenine varied from 1.6 to 1.8, and the 5-methylcytosine content varied from 0.1 to 0.4% . In contrast, the range in percentage of N6-methyladenine and 5-methylcytosine found in the hosts was 0.7 to 2.4 and 0.0 to 1.1, respectively. J Immunol, 1979 Mar, 122(3), 932 - 5 Lack of responsiveness of C3H/HeJ macrophages to lipopolysaccharide: the cellular basis of LPS-stimulated metabolism; Ryan JL et al.; The rate of glucose utilization has been used as a measure of LPS-induced activation of cultures of C3H/HeN and C3H/HeJ spleen cells, peritoneal cells, and purified peritoneal adherent cells . Peritoneal cells utilized 40 to 60 times more glucose than did spleen cells and purified adherent monolayers were more active than mixed peritoneal cells, suggesting that only macrophage metabolism was being measured . The cell preparations for C3H/HeJ mice were not activated by Escherichia coli K235 LPS prepared by extensive phenol extraction, whereas C3H/HeN cells were activated by the LPS . Cells from both strains were activated by a commercially obtained E . coli 0111:B4 LPS and butanol-extracted K235 LPS . The addition of 10% C3H/HeN spleen cells to C3H/HeJ peritoneal cells resulted in a marked enhancement of glucose utilization . These findings suggest that LPS-induced enhancement of macrophage metabolism occurs both by direct action of LPS on macrophages as well as indirectly through activated lymphocytes. Urology, 1979 Mar, 13(3), 321 - 3 Bilateral renal malakoplakia; Ho KL et al.; A case of malakoplakia is presented which was confined to the renal parenchyma of both kidneys in a thirty-five-year-old woman in whom fatal renal failure developed in three weeks . Bilateral renal malakoplakia is rare, and unlike malakoplakia of the bladder, it behaves as a progressive, destructive, and fatal disease. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1363 - 7 Strand exchange in site-specific recombination; Enquist LW et al.; The site-specific recombination system of phage lambda promotes crossovers at its attachment site (att) . In this report we show that when phage are crossed in conditions where only the site-specific recombination system is active, a low frequency of crossovers can also be detected in a region that is close to but does not contain att . These crossovers require the phage int gene, the host hip gene, and the integrity of att . They are not detected if one of the parents carries a substitution of a heterologous attachment site (attB instead of attP) . To explain these findings we suggest that site-specific recombination can proceed by exchange of single strands between the participating chromosomes at att and migration of the resulting junction outside of att. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1213 - 7 Requirements of acetyl phosphate for the binding protein-dependent transport systems in Escherichia coli; Hong JS et al.; In Escherichia coli, acetyl phosphate can be formed from acetyl-CoA via the phosphotransacetylase (phosphate acetyltransferase; acetyl-CoA:orthophosphate acetyltransferase, EC 2.3.1.8) reaction and from acetate (plus ATP) via the acetate kinase (ATP:acetate phosphotransferase, EC 2.7.2.1) reaction . By restricting acetyl phosphate formation to the phosphotransacetylase reaction alone, through the use of metabolic inhibitors, we were able to show that, with pyruvate as a source of energy, mutants defective in phosphotransacetylase are unable to transport glutamine, histidine, and methionine . However, with the same energy source, mutants defective in acetate kinase are normal in the transport of these amino acids . The inability of the phosphotransacetylase mutants to transport is due to their presumed inability to form acetyl phosphate, because pyruvate is found to be metabolized to acetyl-CoA in these mutants . Thus acetyl phosphate has been implicated in active transport . Evidence is also presented that neither the protonmotive force nor the ecf gene product is required for the shock-sensitive transport systems. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1122 - 5 Mutations reducing the activity of c17, a promoter of phage lambda formed by a tandem duplication; Mozola MA et al.; We report the isolation of four independently selected mutations (scs) in the c17 promoter of phage lambda that reduce or eliminate the promoter activity . The c17 promoter is not normally present in lambda, and has been shown to be generated by a tandem duplication which creates a "Pribnow Box," a heptamer sequence implicated in promoter activity . This sequence is located upstream from the site of transcription initiation and is present, with some variation, in all promoters whose sequences have been determined . Analysis of the c17 duplications carrying the scs mutations reveals that three of these mutants carry single base-pair changes in the most highly conserved base pairs of the Pribnow Box and that the other mutation is a reversion to the wild type sequence in this region (i.e., a loss of the duplicated base pairs). Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1089 - 93 Enzymatic insertion of purine bases into depurinated DNA in vitro; Livneh Z et al.; An enzymatic activity that inserts purines into depurinated DNA was found in a soluble enzyme extract of Escherichia coli . This activity brings about the insertion of adenine and guanine into the appropriate apurinic sites in double-stranded DNA by using the corresponding deoxyribonucleoside triphosphates as the purine donors . Magnesium ions are required for this activity, it is inhibited by caffeine, and it does not act on depurinated single-stranded DNA . The insertion activity described here may represent a step in a repair mechanism, "base-insertion repair," whereby apurinic sites (which may occur in double-stranded DNA either due to the removal of damaged purines with specific glycosylases or by spontaneous depurination) are directly filled with the correct missing purine base. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1084 - 8 Biochemical assay designed to detect formation of recombination intermediates in vitro; Potter H et al.; A biochemical assay that is designed to detect recombination intermediates formed in vitro is described . The assay measures the fusion of two essentially homologous plasmids, one of which is radioactively labeled and the other of which carries several copies of the lac operator . The fusion product is radioactive and can be bound to a nitrocellulose filter by lac repressor . This assay for genome fusion is rapid and readily applicable to the many fractions that result during enzyme purification . The fused product is not destroyed in the assay and may be recovered from the filter for further analysis by electron microscopy . The product is then seen to consist of figure 8 structures that can be cleaved by the restriction enzyme EcoRI to give chi forms, structures similar to those recovered from recombination-proficient cells . It is expected that this assay will be useful in the purification of the "recombinase-type" activity detected in crude cell lysates . To demonstrate this point, the assay was applied to the protein fractions recovered from a molecular sieve column . The results indicate that the fusion activity has an apparent molecular weight of 50,000--100,000. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1054 - 8 Localization of the decoding region on the 30S Escherichia coli ribosomal subunit by affinity immunoelectron microscopy; Keren-Zur M et al.; The decoding region of the Escherichia coli ribosome has been localized by affinity immunoelectron microscopy . Valyl-tRNA1Val, derivatized at the alpha-amino end with the dinitrophenyl group, was bound to the ribosomal P site of 70S ribosomes and crosslinked specifically to 16S RNA by 310- to 325-nm irradiation . Previous work has shown that crosslink occurs between the 5' anticodon base of the tRNA and a pyrimidine in the 3' one-third of the 16S RNA . By reaction of the dinitrophenyl group with antibody, dimers of the 30S tRNA covalent complexes were prepared containing one covalently attached tRNA per 30S subunit . Electron microscopic visualization of the antibody attached to the dinitrophenyl group located the position of the 3' end of the tRNA . Several sites, with a strong preference for the large protrusion or cleft region in the upper one-third of the subunit, were found . The multiplicity of sites is likely due to the freedom of orientation of the 3' end of the tRNA which is approximately 80 A from the site of crosslinking . By extrapolating this distance from each of the antigenic sites, we concluded that the anticodon of tRNA when in the P site is probably located in the cleft region of the 30S subunit. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1040 - 4 Interaction of Escherichia coli ribosomal protein S1 with ribosomes; Draper DE et al.; The binding affinity of Escherichia coli ribosomal protein S1 for 30S ribosomal particles has been determined by a sucrose gradient band sedimentation technique; the association constant (K) for the binding of one S1 protein per active 30S ribosomal subunit is approximately 2 X 10(8) M-1 . The involvement of the two polynucleotide binding sites of S1 protein (site I binding single-stranded DNA or RNA, and site II binding single-stranded RNA only) in the S1--ribosomal interaction have been examined by competition experiments with polynucleotides of known affinity for the two sites . We find that site I does not contribute to the interaction; site II binding appears to provide a major part of the binding free energy, presumably by interaction of S1 with the 16S rRNA of the 30S particle . The remaining binding free energy is probably derived from the interaction of S1 protein with other proteins of the 30S subunit . The affinity of S1 for 70S ribosomes is about the same as that for the 30S subunit; the affinity of S1 for 50S subunits is much less . Binding affinities and stoichiometries of S1 protein with "inactive" 30S ribosomal subunits have also been examined. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1035 - 9 High-frequency transformation of yeast: autonomous replication of hybrid DNA molecules; Struhl K et al.; A set of vector DNAs (Y vectors) useful for the cloning of DNA fragments in Saccharomyces cerevisiae (yeast) and in Escherichia coli are characterized . With these vectors, three modes of yeast transformation are defined . (i) Vectors containing yeast chromosomal DNA sequences (YIp1, YIp5) transform yeast cells at low frequency (1--10 colonies per microgram) and integrate into the genome by homologous recombination; this recombination is reversible . (ii) Hybrids containing endogenous yeast plasmid DNA sequences (YEp2, YEp6) transform yeast cells at much higher frequency (5000--20,000 colonies per microgram) . Such molecules replicate autonomously with an average copy number of 5--10 covalently closed circles per yeast cell and also replicate as a chromosomally integrated structure . This DNA may be physically isolated in intact form from either yeast or E . coli and used to transform either organism at high frequency . (iii) Vectors containing a 1.4-kilobase yeast DNA fragment that includes the centromere linked trp1 gene (YRp7) transform yeast with an efficiency of 500--5000 colonies per microgram; such molecules behave as minichromosomes because they replicate autonomously but do not integrate into the genome . The uses of Y vectors for the following genetic manipulations in yeast are discussed: isolation of genes; construction of haploid strains that are merodiploid for a particular DNA sequence; and directed alterations of the yeast genome . General methods for the selection and the analysis of these events are presented. Nucleic Acids Res, 1979 Mar, 6(3), 915 - 30 Construction and analysis of recombinant DNAs containing a structural gene for rat prolactin; Gubbins EJ et al.; Poly A-containing RNA enriched in prolactin-coding sequences was isolated from female rate pituitaries after induction with diethylstilbesterol . Double stranded cDNA was synthesized from this RNA and inserted into plasmid pBR322 at the Pst I site via the poly(dG):polyy(dC) tailing method . E . coli was transformed with this DNA and the recombinant plasmid in one of the transformants characterized in detail . About half of its 900 base pair cDNA insert was sequenced . The DNA sequence is consistent with most of the reported amino acid sequence of rat preprolactin . In addition, the recombinant plasmids in two of the other transformants appear to contain growth hormone coding sequences. Nucleic Acids Res, 1979 Mar, 6(3), 899 - 914 Chemical modification study of aminoacyl-tRNA conformation; Negishi K et al.; Chemical reactivity of cytosines in 32P-labeled E . coli tRNA1Leu, E . coli tRNAPhe and yeast tRNAPhe before and after aminoacylation was examined by use of a cytosine-specific reagent, semicarbazide-bisulfite mixture . In all the three tRNA species examined, the cytosine residues that were susceptible to the modification were the same in the aminoacylated tRNA and the unacylated tRNA . Only a limited number of the cytosine residues were modifiable: those that occur in the anticodon, the 3'-CCA terminus, the D-loop, and the extra loop . The sites accessible by the reagent are in good agreement with the general three-dimensional structure of tRNA proposed in literature . These results indicate that the gross conformation of these tRNAs does not change on aminoacylation, and consequently favor the view that the T psi C(G) sequence could become exposed in later steps of protein synthesis in order to achieve the binding of aminoacyl tRNA to ribosomes. Nucleic Acids Res, 1979 Mar, 6(3), 849 - 65 Cloning, mapping and expression of the genetic determinant that encodes for the K88ab antigen; Mooi FR et al.; The K88 antigen, a plasmid-specified virulence factor of E . coli involved in porcine neonatal diarrhoea, is often found to be associated with the ability to metabolize raffinose (Raf) . Plasmid pRI8801 (51 megadalton) was used to clone the determinants of K88 and Raf with the vector pBR322 . K88 was found to be encoded by a 7.7 megadalton HindIII fragment . The expression was highly dependent on the orientation of the HindIII fragment within pBR322 . By in vitro generation of deletions, the HindIII fragment was reduced in size to 4.3 megadalton . The expression of K88 by pRI8801 and the recombinant plasmids was studied using an enzyme-linked immunosorbent assay . Raf was found to be located on a 4.0 megadalton SalI fragment . A physical map of pRI8801 was constructed . The K88 antigen and Raf genes are not closely linked but separated by a stretch of DNA of about 20 megadalton. Nucleic Acids Res, 1979 Mar, 6(3), 1177 - 87 Photosensitized formation of thymine dimers in DNA by tyramine, tyrosine and tyrosine-containing peptides; Kaneko M et al.; The formation of Thy-Thy in DNA in the presence of tyramine, tyrosine and tyrosine-containing peptides such as Lys-Tyr and Lys-Tyr-Lys was studied with monochromatic UV irradiation . The formation of Thy-Thy by UV irradiation was enhanced in the presence of these compounds . The action spectrum of the photosensitization has a peak near 280 nm corresponding to the absorption spectrum of tyrosine . The triplet quencher reduced the sensitization substantially . The sensitization in native DNA was more than six times larger than that in denatured DNA . increasing the concentration of salts suppressed the sensitization . The nature of the interaction between DNA and the sensitizer is discussed. Nucleic Acids Res, 1979 Mar, 6(3), 1123 - 33 In vitro methylation of yeast tRNAAsp by rat brain cortical tRNA-(adenine-1) methyltransferase; Salas CE et al.; Rat brain cortices from young animals contain large amounts of tRNA (adenine-1)methyltransferase(s) . The enzyme(s) can methylate E . coli tRNA and to a lower degree yeast tRNA . Among yeast tRNA species which can be methylated we have selected tRNAAsp as a substrate for the brain enzyme . The digestions of in vitro methylated {Me-3H}-tRNAAsp with pancreatic and/or T1 ribonucleases followed by chromatographies on DEAE-cellulose, 7 M urea, suggested that the methylation of tRNAAsp occurred at a single position within the D-loop . Further digestion of the radioactive oligonucleotide recovered after DEAE-cellulose chromatography by phosphomonoesterase and snake venom phosphodiesterase enzymes followed by bidimensional thin layer chromatography enabled us to determine the location of the adenine residue which becomes methylated by the brain enzyme . This one resulted to be the adenine 14 in the D-loop of yeast tRNAAsp. Nucleic Acids Res, 1979 Mar, 6(3), 1111 - 22 Nucleotide sequence of the transposable DNA-element IS2; Ghosal D et al.; The complete sequence of the transposable DNA element IS2 in gal OP-308:: IS2 (I) has been determined . This element is 1.327 bp long . The integrated element is flanked by a five base pair long sequence duplication . The termini of IS2 are not perfect inverted repeats, but a close approximation. Mol Biol (Mosk), 1979 Mar-Apr, 13(2), 388 - 401 {Modification of the RNA-polymerase of Escherichia coli by diethylpyrocarbonate}; Rozovskaia TA et al.; E . coli DNA dependent RNA polymerase was modified by diethylpyrocarbonate . Optical and kinetic properties of the reaction were studied . More than 90% of RNA polymerase activity is inhibited by introduction of 9--11 ethoxyformyl groups per enzyme molecule without loss of its ability to bind DNA template . Furthermore the modified enzyme is able to form tight complexes with DNA and to compete with native enzyme for the formation of rifampicin-resistant complex . The ratio of the complex formation constants for the native and modified enzyme was determined to be equal to 10 . The enzyme modified to such extent loses the activity in DNA dependent RNA as well as pppApU synthesis . Vmax value rather than Km value for both ATP and UTP decreases following the modification reaction . Incubation of the enzyme modified to the 10% of residual activity with 0.2 M hydroxylamine for 2 hours results in restoration of RNA polymerase activity . Most but not all of the modified histidyl residues restore their native structure . Two of 13 histidyl residues were modified irreversibly due to Bamberger's cleavage reaction, but these two residues were found to be unessential for RNA polymerase activity . Reaction with higher concentration of the diethylpyrocarbonate induces modification of more than 15--20 histidyl residues and leads to irreversible inactivation of the enzyme . Nevertheless the modification of the additional histidyl redidues was reversible as well as the modification of the first 11 residues . RNA polymerase modified to such extent loses the ability to bind DNA . Preformation of the initiated ternary complex of RNA polymerase with template and product fails to protect the enzyme from reversible inactivation at a low reagent concentration, but markedly decreases the rate of the irreversible and unspecific modification of sulfhydryl or amino groups of the enzyme . Reaction with the ternary complex results in reversible inactivation of the enzyme with respect to elongation of RNA chains as well as the pyrophosphate exchange reaction . The complex itself was, however, completely stable under the reaction conditions and the enzyme subunit structure was also conserved after the reaction . Evidently, the mild modification of the histidyl residues with diethylpyrocarbonate selectively inhibits RNA chain elongation. Mol Biol (Mosk), 1979 Mar-Apr, 13(2), 281 - 91 {Isolation and properties of DNA-cytosine methyltransferase from Escherichia coli C}; Bogdarina IG et al.; The method of isolation and partial purification of DNA-cytosine-methyltransferase (DC-methylase) from E . coli C is described . The enzyme underwent approximately 100-fold purification . The obtained preparation of DC-methylase can be additionally considerably purified by sedimentation in sucrose gradient . Native molecular weight of DC-methylase from E . coli C . is 70,000 . The activity of enzyme does not depend on the Mg2+ ions . DC-methylase E . coli C provides DNA of lambda phage in vitro with full resistance against restriction endonuclease EcoRII . In DNA methylated by DC-methylase the modified cytosine, mainly in C-MC and C-MC-T sequences, corresponds to the pyrimidine sequences of specific site EcoRII . DNA of lambda.B phage contains approximately 80 sites for modification by DC-methylase E . coli C . The results obtained point to the same specificity in vitro of DNA-cytosine-methylase E . coli C and DNA-methylase EcoRII. Mikrobiologiia, 1979 Mar-Apr, 48(2), 307 - 10 {Morphological changes in the cells of Escherichia coli damaged by nitrous yperite}; Lobanok EV et al.; Nitrous yperite supressed division of E . coli cells; as a result, filaments and giant abnormal cells appeared . Apparently, giant forms were caused by defects in the cell wall induced by nitrous ypertie due to a mutation in the mon gene of the strain . These cells gave rise to normal cells as well as to small bacteria resembling mini-cell. J Bacteriol, 1979 Mar, 137(3), 1439 - 42 Enhancement of deoxyribonucleic acid polymerase I-directed repair synthesis in toluene-treated Escherichia coli after growth in the presence of low levels of N-methyl-N'-nitro-N-nitrosoguanidine; Billen D et al.; Deoxyribonucleic acid polymerase I-directed repair synthesis can be selectively measured in toluene-treated Escherichia coli cells exposed to alkylating chemicals such as N-methyl-N'-nitro-N-nitrosoguanidine . Prior growth of the cells in the presence of a low dose of N-methyl-N'-nitro-N-nitrosoguanidine results in an enhanced deoxyribonucleic acid polymerase I-directed repair synthesis and an increase in single-strand breaks. J Bacteriol, 1979 Mar, 137(3), 1421 - 4 Enumeration and identification of IS3 elements in Escherichia coli strains; Deonier RC et al.; Escherichia coli K-12 strains ordinarily contain five IS3 elements . Three of these correspond to previously mapped IS3 elements (R . C . Deonier, G . R . Oh, and M . Hu, J . Bacteriol . 129:1129--1140, 1977; S . Hu, E . Ohtsubo, and N . Davidson, J . Bacteriol . 122:749--763, 1975), and two additional IS3 elements are identified . The distribution of IS3 elements among deoxyribonucleic acid fragments generated by digestion with EcoRI indicates a basic pattern from which deviation is detected. J Bacteriol, 1979 Mar, 137(3), 1243 - 52 Enzymatic degradation of uracil-containing deoxyribonucleic acid . V . Survival of Escherichia coli and coliphages treated with sodium bisulfite; Simmons RR et al.; A number of mutants of Escherichia coli defective in the ung gene (structural gene for uracil-deoxyribonucleic acid {ura-DNA} glycosylase) are shown to be abnormally sensitive to treatment with sodium bisulfite when compared with congenic ung+ strains . These results provide further evidence that sodium bisulfite causes the deamination of cytosine to uracil in DNA and that ura-DNA glycosylase is required for the repair of U-G mispairs . The effect of the chemical is apparently selective with respect to base damage; coliphages containing cytosine in their DNA are inactivated by treatment with sodium bisulfite, whereas those containing hydroxymethylcytosine are not . ura-DNA glycosylase and the major apurinic-apyrimidinic endonuclease of E . coli may function in the same repair pathway, since the extent of inactivation of a congenic set of strains which are ung xth (structural gene for the major apurinic-apyrimidinic endonuclease of E . coli) or ung xth+ is the same. J Bacteriol, 1979 Mar, 137(3), 1151 - 7 Chromosome replication and cell division in plasmid-containing Escherichia coli B/r; Weinberger M et al.; The kinetics of chromosome replication and cell division have been examined in recA mutants of Escherichia coli B/r containing F' plasmids of various sizes . Plasmid-mediated alterations in growth properties were detected only with the presence of the larger F' plasmids, and were reflected in decreased mean cell sizes and growth rates . The lengths of C and D in all plasmid-containing strains were in accord with the values for plasmid-free parental strains growing with similar generations times . The findings were consistent with an absence of competition between the chromosomal and extrachromosomal replicons for rate-limiting components involved in the initiation of deoxyribonucleic acid synthesis or in the elongation of deoxyribonucleic acid chains. J Bacteriol, 1979 Mar, 137(3), 1111 - 8 Isolation and properties of Escherichia coli K-12 mutants impaired in the utilization of gamma-aminobutyrate; Metzer E et al.; We have isolated mutants of Escherichia coli K-12 CS101B that have lost the ability to utilize gamma-aminobutyrate as a source of nitrogen . One class of mutants, which were not affected in the utilization of other nitrogen sources (proline, arginine, glycine), included many isolates with lesions in gamma-aminobutyrate transport or in its transamination and one mutant completely devoid of succinic semialdehyde dehydrogenase activity and exhibiting low gamma-aminobutyrate transport and transamination . gamma-Aminobutyrate-utilizing revertants of the latter recovered full transport and transamination capacities but remained dehydrogenaseless . Another class of mutants showed pleiotropic defects in nitrogen metabolism . One such mutant was lacking glutamate synthase activity . The genes specifying the synthesis of gamma-aminobutyrate permease, gabP, gamma-aminobutyrate transaminase, gabT, and succinic semialdehyde dehydrogenase, gabD, and the control gene, gabC, that coordinately regulates their expression all form a cluster on the E . coli chromosome, linked to the srl and recA loci (at 57.5 min) . The mutations with pleiotropic effects on the metabolism of nitrogenous compounds are not linked to the gab cluster. J Bacteriol, 1979 Mar, 137(3), 1088 - 94 Identification of the ftsA gene product; Lutkenhaus JF et al.; A nonsense mutation was identified in the essential cell division gene ftsA of Escherichia coli . A gamma-transducing phage was isolated which complemented this mutation . This phage programmed the synthesis of four bacterial proteins in UV-irradiated cells . By substituting the nonsense mutation for the ftsA+ allele in this transducing phage and comparing the proteins programmed by it in UV-treated Su+ and Su- cells, the product of the ftsA gene was identified as a protein with a molecular weight of 50,000. J Bacteriol, 1979 Mar, 137(3), 1084 - 7 Occurrence of 1-methyladenosine and absence of ribothymidine in transfer ribonucleic acid of Mycobacterium smegmatis; Vani BR et al.; The minor base composition of Mycobacterium smegmatis tRNA has been studied . Thin-layer chromatographic patterns of a ribonuclease T2 digest of mycobacterial tRNA indicated the presence of appreciable amounts of 1-methyladenosine (which is commonly present only in eucaryotic tRNA), dihydrouridine, and 7-methylguanosine . Ribothymidine was absent . The S-adenosylmethionine-dependent tRNA methylases of M . smegmatis catalyzed the formation of 1-methyladenosine when Escherichia coli tRNA was used as acceptor . Similarly, E . coli extracts methylated the tRNA of M . smegmatis, forming ribothymidine. Mutat Res, 1979 Mar, 60(1), 13 - 23 Mutation induction by thymidine deprivation in Escherichia coli B/r . I . Influence of caffeine; Mennigmann HD et al.; The addition of caffeine to the plating medium after thymine deprivation of E . coli WP2 uvr+ thyA or WP2 uvrA thyA had no influence on survival . Caffeine, however, reduced the frequency of mutants . The hypothesis is presented that the reduced mutagenesis is due to the sensitivity to caffeine of an inducible error-prone repair mechanism operating during thymine deprivation and after the re-addition of thymine. J Exp Med, 1979 Mar 1, 149(3), 669 - 85 Form variation in Escherichia coli K1: determined by O-acetylation of the capsular polysaccharide; Orskov F et al.; The chemical basis for the alternating antigenic change called form variation noted for the Escherichia coli K1-capsular polysaccharide has been shown by 13C nuclear magnetic resonance to be a result of random O-acetylation of C7 and C9 carbons of the alpha-2-8-linked sialic acid homopolymer . A serologic method (antiserum agar) was developed to identify and isolate the form variants . The O-acetyl positive and O-acetyl negative K1 polysaccharides had unique biochemical and immunologic properties . The O-acetyl-positive variants resisted neuraminidase hydrolysis in contrast to the susceptibility of the O-acetyl negative variant to this enzyme . In addition, O-acetylation altered the antigenicity of the O-acetyl polysaccharides . When injected as whole organisms, O-acetyl positive organisms produced anti-K1 -antibodies in rabbits specific for this polysaccharide variant . O-acetyl negative organisms were comparatively less immunogenic; however, antibodies induced by these organisms reacted with both K1 polysaccharide variants . Burros, injected with either variant, produced antibodies reactive with both K1 polysaccharides. Eur J Biochem, 1979 Mar, 94(2), 477 - 84 Translational fidelity and specificity of ribosomes cleaved by cloacin DF13; Twilt JC et al.; The effect of cloacin DF13 cleavage on several functional properties of the ribosome has been studied in a translational system in vitro . Cleaved ribosomes synthesize relatively shorter polypeptide chains on synthetic and natural templates . Their translational specificity is, however, unchanged as judged by the read-out of MS2 RNA . Here, cleaved as well as control ribosomes start translation only on the coat cistron of the phage RNA . Cloacin cleavage of ribosomes increases their fidelity of translation . Differential inhibition of translation of synthetic and natural template was not observed. Surg Gynecol Obstet, 1979 Mar, 148(3), 361 - 6 Methylprednisolone in the prevention of cerebral hemodynamic and metabolic disorders during endotoxin shock in the dog; Emerson TE Jr et al.; These results provide evidence that steroid pretreatment and subsequent post-treatment prevent cerebral hemodynamic and metabolic alterations during four hours of Escherichia coli endotoxin shock in the dog . However, in this study, no data are provided on how the steroid prevents an increase in cerebral vascular resistance, and no clear answer is available in the literature . While active vasodilation or alpha-adrenergic blocking properties, or both, have been attributed to glucocorticoids, recent evidence does not support these findings during normal conditions or circulatory shock . If the increase in cerebral vascular resistance is passive, steroids may help by preventing platelet aggregation, cell disruption and subsequent microvascular plugging . Intravenously administered fluids, dextran-saline solution, while in themselves are probably not important to survival, may augment cerebral blood flow during shock through a blood dilutional effect . Finally, it is possible that steroids act to permit normal, long term cerebral auto-regulation, which is apparently impaired during endotoxin shock in the dog. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1194 - 8 Subcellular localization of acyl carrier protein in leaf protoplasts of Spinacia oleracea; Ohlrogge JB et al.; This communication demonstrates that all de novo fatty acid biosynthesis in spinach leaf cells requires acyl carrier protein (ACP) and occurs specifically in the chloroplasts . Antibodies raised to purified spinach ACP inhibited at least 98% of malonyl CoA-dependent fatty acid synthesis by spinach leaf homogenates . Therefore, the presence of ACP in a compartment of the spinach leaf cell would serve as a marker for de novo fatty acid biosynthesis . A radioimmunoassay capable of detecting 10(15) mol (10(-11) g) of spinach ACP was developed to measure the levels of ACP in leaf cell components isolated by sucrose gradient centrifugation of a gentle lysate of spinach leaf protoplasts . All of the ACP of the leaf cell could be attributed to the chloroplast . Less than 1% of the ACP associated with chloroplasts resulted from binding of free ACP to chloroplasts . Of interest, ACP from Escherichia coli, soybean, and sunflower showed only partial crossreactivity with spinach ACP by the radioimmunoassay . These results strongly suggest that, in the leaf cell, chloroplasts are the sole site for the de novo synthesis of C16 and C18 fatty acids . These fatty acids are then transported into the cytoplasm for further modification and are either inserted into extrachloroplastic membrane lipids or returned to the chloroplast for insertion into lamellar membrane lipids. Rev Infect Dis, 1979 Mar-Apr, 1(2), 357 - 69 Superoxide, hydrogen peroxide, and oxygen tolerance of oxygen-sensitive mutants of Escherichia coli; Hassan HM et al.; Oxygen-intolerant mutants of Escherichia coli K12 were selected by a replica plating technique after treatment with the mutagen, N-methyl-N'-nitro-N-nitrosoguanidine, to a lethality of 99.5% . One group of mutants had lost the ability to induce both peroxidase and catalase when exposed to oxygen but retained the ability to induce the manganese-superoxide dismutase . The second group of mutants had lost the ability to induce the activity of all these enzymes . Failure to induce peroxidase and catalase was associated with enhanced susceptibility of the bacteria to the lethal effect of oxygen . When a member of the first group of mutants was prevented from producing the manganese-superoxide dismutase by the presence of puromycin, its susceptibility to the lethal effects of oxygen was greatly increased . Two types of revertants were seen . In one group the ability to induce enzyme activity was recovered and was accompanied by the return of oxygen tolerance . Members of the other group lost the ability to respire and, therefore, no longer produced O2- AND H2O2 . These results indicated that enzymic scavenging of both H2O2 and O2- provides an important defense against oxygen toxicity . The parallel loss of peroxidase and catalase, which was seen in all mutants, suggests that these enzymes constitute a precursor-product pair in E . coli . The parallel loss in two of these mutants of peroxidase, catalase, and the manganese-superoxide dismutase suggests a control linkage for these enzymes, the basis of which remains to be explored. Eur J Biochem, 1979 Mar, 94(2), 445 - 50 Replication of M13 duplex DNA in soluble extracts of Escherichia coli . Effect of helix-destabilising proteins; van Dorp B et al.; Soluble extracts of M13-am5-infected Escherichia coli cells can carry out multiple rounds of M13 duplex DNA replication when supplemented with helix-destabilising protein of E . coli . Similarly addition of the helix-destabilising M13 gene 5 protein in low concentrations (up to 30 micrograms/ml) stimulates the replication of double-stranded M13 DNA . In contrast, higher concentrations of gene 5 protein (but not of E . coli helix-destabilising protein) cause a preferential inhibition of complementary strand synthesis resulting in a switch from double-strand replication to single-strand synthesis . Depending on the addition of the appropriate amounts of these two helix-destabilising proteins either stage of M13 DNA replication can now be studied with cell-free preparations. Z Naturforsch {C}, 1979 Mar-Apr, 34(3-4), 248 - 52 On the dynamic model of tRNA: recent experimental findings; Gurel O; Recent experimental findings are shown to support various aspects of the dynamic model of tRNA proposed earlier and the interaction between tRNA, mRNA and the ribosomal RNA's . Extra loop of tRNA molecule is suggested to play a role in recognizing the corresponding amino acids and a correlation is presented between the tRNA molecules and the corresponding amino acids as tabulated by the genetic tableau. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1126 - 30 Specialized transducing phage lambda carrying the genes for coupling factor of oxidative phosphorylation of Escherichia coli: increased synthesis of coupling factor on induction of prophage lambda asn; Kanazawa H et al.; Studies were made of the synthesis of the coupling factor complex (F1--F0) of oxidative phosphorylation after prophage induction of a set of Escherichia coli strains lysogenic for defective transducing phage lambda asn, lambda uncA, or lambda bglC . The transducing phages had been isolated from a strain of E . coli carrying prophage lambda cI857 S7 within the bglB gene located near the unc gene cluster {Miki, T., Hiraga, S., Nagata, T . & Yura, T . (1978) Proc . Natl . Acad . Sci . USA 75, 5099--5103} . When lysogenic cells carrying lambda asn and lambda cI857 S7 were induced at high temperature, synthesis of the F1-ATPase portion of the complex increased to severalfold that of the noninduced cells . In contrast, no increase was observed upon thermoinduction of cells carrying lambda uncA or lambda bglC . The number of membrane sites that could bind purified F1-ATPase also increased significantly upon induction by lambda asn but not by lambda uncA or lambda bglC . In addition, F1-depleted membranes prepared from lambda asn-induced bacteria required more dicyclohexylcarbodiimide to seal the proton pathway than did those from noninduced bacteria . These results strongly suggest that lambda asn carries a set of bacterial genes coding for all the F1 polypeptides (the alpha, beta, gamma, delta, and probably the epsilon subunits) and at least some of the genes involved in formation of F0 polypeptides . Although lambda uncA carries the structural gene (uncA) for the alpha subunit of F1-ATPase, it apparently does not carry the whole set of F1--F0 genes. J Bacteriol, 1979 Mar, 137(3), 1219 - 26 Arrangement of the genes coding for ribosomal ribonucleic acids in Neurospora crassa; Free SJ et al.; We have cloned and characterized Neurospora crassa ribosomal deoxyribonucleic acid (rDNA) . The rDNA is found as a tandemly repeated 6.0-megadalton sequence . We have mapped a portion of the rDNA repeat unit with respect to its sites for 13 restriction endonucleases and defined those regions coding for the 5 . 8S, 17S, and 26S ribosomal ribonucleic acids (rRNA's) . We have also isolated several clones containing 5S rRNA sequences . The 5S rRNA coding sequences are not found within the rDNA repeat unit . We found that the sequences surrounding the 5S rRNA coding regions are highly heterogeneous. Infect Immun, 1979 Mar, 23(3), 700 - 5 Behavior of Escherichia coli K antigens K88ab, K88ac, and K88ad in immunoelectrophoresis, double diffusion, and hemagglutination; Guinee PA et al.; Porcine enteropathogenic Escherichia coli strains were found to possess a variant of the K88 antigen provisionally termed K88ad . We propose to include this antigen into the international E . coli typing scheme . Ultrasonic extracts of field strains of E . coli possessing the K88ab, K88ac, or K88ad antigen and their E . coli K-12 K88+ transconjugants showed a specific K88 precipitation line in immunoelectrophoresis and double diffusion only when grown at 37 degrees C, but not when grown at 18 degrees C . By using agarose gels, K88ab, K88ac, and K88ad antigens showed anodic mobility in immunoelectrophoresis . When using Difco Noble agar gels, K88ad was not mobile or anodic, K88ab was cathodic; K88ac of 17 strains was cathodic and of 24 strains was anodic . The immunoelectrophoretic behavior of a K88 antigen (K88ab, K88ac, or K88ad) did not alter after transfer of the corresponding plasmid to E . coli K-12 . Anodic and cathodic K88ac antigens could not be distinguished serologically . The differences between the results obtained in Noble agar gels and agarose gels are due to electro-endosmotic flow . We describe a procedure which increases the detection level of K88+ transconjugants in a mating mixture . It is based on the specific mannose-resistant attachment of K88+ cells to guinea pig erythrocytes. Can J Microbiol, 1979 Mar, 25(3), 335 - 9 Response of various cell lines to Escherichia coli toxic products; Konowalchuk J et al.; Seven cell lines were compared with Y-1 and Vero cells for morphological response to E . coli toxic products . FL and WI-38, like Y-1 and Vero, responded to heat-labile enterotoxin . FL, HEp-2, HeLa, and BGM, like Vero but not Y-1, responded to heat-labile cytotoxin . FL was comparable to Vero in sensitivity to these two toxic products. Proc Natl Acad Sci U S A, 1979 Mar, 76(3), 1099 - 103 Escherichia coli adenylate cyclase complex: regulation by the proton electrochemical gradient; Peterkofsky A et al.; Sugars such as glucose are transported into Escherichia coli by a coupled phosphorylation mechanism (the phosphoenolpyruvate:sugar phosphotransferase system, PTS) . Transport of sugars through the PTS results in inhibition of adenylate cyclase {ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1} activity by a mechanism involving a change in the state of phosphorylation of PTS proteins . Other sugars (e.g., lactose) are transported without modification by a mechanism involving proton cotransport, which requires a proton motive force across the cell membrane . We show here that uptake of sugars through the lactose transport system results in inhibition of adenylate cyclase activity if the proton symport mechanism is also active . The protonophore carbonyl cyanide m-chlorophenylhydrazone also inhibits adenylate cyclase activity . These data suggest that the steady-state electrochemical proton gradient regulates the activity of adenylate cyclase . We propose that sugar-dependent inhibition of adenylate cyclase activity may occur by either of two mechanisms . Sugars transported by the PTS inhibited adenylate cyclase activity by dephosphorylation of a regulatory protein, while sugars transported by the proton motive force system inhibit adenylate cyclase activity as a result of collapse of the proton electrochemical gradient. Biol Bull Acad Sci USSR, 1979 Mar-Apr, 6(2), 172 - 8 Effect of beta-indoleacetic acid on RNA and protein synthesis in Escherichia coli; Drobysheva NA et al.; The effect of different concentrations of beta-indoleacetic acid (IAA) on RNA and protein synthesis and the stability of associations of E . coli ribosomes at 3 mM Mg2+ was investigated . IAA in a concentration of 100 microgram/ml had a slight stimulatory effect on the incorporation of {14C}leucine into proteins and stimulated the incorporation of {14C}uracil into the RNA of intact cells more noticeably . In the presence of 2500 microgram/ml of the auxin RNA synthesis was inhibited by 65% and protein synthesis by 40-50% as compared to the control . In a cell-free system containing poly(U) IAA in concentrations of 10-100 microgram/ml increased polyphenylalanine synthesis by an average of 30%, while at high concentrations IAA noticeably inhibited its synthesis . It was found that the proportion of stable ribosomes in lysates obtained from cells incubated with IAA in concentrations of 250 and 1000 microgram/ml decreased to 18 and 3% . It is suggested that the inhibition of protein synthesis in E . coli by IAA is due to inhibition of the initiation of translation. Biochim Biophys Acta, 1979 Feb 27, 561(2), 396 - 402 Utilization in vitro of deoxyuridine triphosphate in DNA synthesis by DNA polymerases alpha and beta from calf thymus; Yoshida S et al.; DNA polymerases alpha and beta (EC 2.7.7.7.) from calf thymus could utilize dUTP as a substrate for DNA synthesis as well as DNA polymerase I of Escherichia coli . Deoxyuridylate was incorporated into DNA by replacing deoxythymidylate and supported the further elongation of DNA chains on activated DNA or on the intiated homopolymers, poly(dA) . (dT)10 and poly(rA) . (dT)10 . The rate of the incorporation of deoxyuridylate into DNA varied from 50 to 160% of that of deoxythymidylate, depending on the nature of the template primers and the species of DNA polymerase used . The apparent Km values for dUTP were very similar to those for dTTP . Uracil DNA-glycosylase excised efficiently the uracil residues in products of DNA polymerase reactions with either activated calf thymus DNA or initiated homopolymers. Biochim Biophys Acta, 1979 Feb 27, 561(2), 435 - 44 A purified nucleoprotein fragment of the 30 S ribosomal subunit of Escherichia coli; Spitnik-Elson P et al.; A '13 S' nucleoprotein fragment was isolated from a nuclease digest of Escherichia coli 30-S ribosomal subunits and purified to gel electrophoretic homogeneity . It contained two polynucleotides, of about 1.1 . 10(5) and 2.5 . 10(4) daltons, which separated when the fragment was deproteinized . The major protein components were S4, S7 and S9/11, with S15, S16, S18, S19 and S20 present in reduced amount. Biochim Biophys Acta, 1979 Feb 27, 561(2), 312 - 23 Genomic integrity of T1 DNA after gamma-and ultraviolet irradiation; Martin-Bertram H et al.; T1 DNA, gamma-irradiated in the phage particle or irradiated with ultraviolet light was checked for structural integrity by kinetics of melting and reannealing . gamma-Irradiated DNA differed in all thermokinetic properties by a factor of 3-4 from DNA degraded by mechanical or enzymatical treatments . Ultraviolet irradiation caused much smaller effects than gamma-irradiation . Considering the frequency of pyrimidine dimers in relation to the gamma-ray induced lesions, strong evidence can be derived, that in addition to single base damages, local denatured regions are produced by gamma-irradiation . Such regions, formed possibly by direct absorption of radiation energy in DNA, i.e . by primary ionizations, are associated with base lesions and are passed over during reannealing. Biochim Biophys Acta, 1979 Feb 27, 561(2), 294 - 300 Escherichia coli Hfr-DNA degradation in endonuclease I-deficient minicells; Khachatourians GG; {3H}Thymidine (dThd)-labelled Hfr DNA was transferred by conjugation into Escherichia coli F- minicells harvested from an endonuclease I-deficient (endI-) strain and its iosgenic wild type (endI+) parent . The susceptibility of this DNA to attack by DNAase was examined . The kinetics of in vivo conversion of {3H}dThd-labelled DNA into acid soluble radioactivity was examined . This activity, attributed to exonuclease action was the same for both strains . Contribution of endonuclease I was measured by an analysis of changes in weight-average (Mw) and number-average (Mn) molecular weight distribution of DNA molecules recovered from minicells . Reduction in Mw was greater in the endI-strain . The ratio Mn/Mw changed drastically during the incubation period of endI- minicells, but remained unchanged in the endI+ strain . These experiments suggest that the presence of the endI- mutation in minicell-producing strain chi1268 leads to a greater loss in M2 of Hfr DNA conjugally transferred into the minicells. Biochim Biophys Acta, 1979 Feb 26, 576(2), 263 - 8 Presence of the epsilon-(gamma-glutamic)lysine crosslink in cellular proteins; Matacic SS et al.; The epsilon-(gamma-glutamic)lysine bond is a covalent interaction which has been found to crosslink polypeptide chains of a number of extracellular proteins . Among known covalent bonds crosslinking protein chains, it is unique in that it is formed directly by enzymatic catalysis, a property which may also endow Glu-Lys crosslink formation with important intracellular functions . We found glutamic-lysine bonds to be present in the procaryote, Escherichia coli, in primitive eucaryotes such as the slime mold, Physarum polycephalum, and the ciliate, Paramecium aurelia, and in muscle cells of a bird and a mammal . Our data show that, although Glu-Lys bonds occur in low concentrations in cellular proteins, they are nevertheless widely distributed . Evidence is also presented indicating that the low levels of the Glu-Lys bonds we measure in the proteins of various cells types are not artifacts of our analytical procedures. Mol Gen Genet, 1979 Feb 26, 170(2), 187 - 9 Methylgroups of ribosomal protein L11 are not related to the synthesis of ppGpp; Rohl R et al.; Ribosomes carrying either a normally methylated or an undermethylated L11, respectively, were tested with respect to the stringency reaction in the presence of crude stringent factor . Systems with either kind of ribosomes synthesize ppGpp with the same efficiency . The ppGpp synthesis in presence of ribosomes with undermethylated L11 depends also on stringent factor, mRNA and deacylated tRNA whereas aminoacyl-tRNA and peptidyl-analogous tRNA show no effect . Thus, the absence of methylation in L11 does not influence the stringency reaction. J Biol Chem, 1979 Feb 25, 254(4), 1408 - 13 Recognition sequence of the dam methylase of Escherichia coli K12 and mode of cleavage of Dpn I endonuclease; Geier GE et al.; The recognition sequence for the dam methylase of Escherichia coli K12 has been determined directly by use of in vivo methylated ColE1 DNA or DNA methylated in vitro with purified enzyme . The methylase recognizes the symmetric tetranucleotide d(pG-A-T-C) and introduces two methyl groups per site in duplex DNA with the product of methylation being 6-methylaminopurine . This work has also demonstrated that Dpn I restriction endonuclease cleaves on the 3' side of the modified adenine within the methylated sequence to yield DNA fragments possessing fully base-paired termini . All sequences in ColE1 DNA methylated by the dam enzyme are subject to double strand cleavage by Dpn I endonuclease . Therefore, this restriction enzyme can be employed for mapping the location of sequences possessing the dam modification. J Biol Chem, 1979 Feb 25, 254(4), 1016 - 21 Biosynthesis in Escherichia coli of sn-glycerol 3-phosphate, a precursor of phospholipid . Palmitoyl-CoA inhibition of the biosynthetic sn-glycerol-3-phosphate dehydrogenase; Edgar JR et al.; Homogeneous biosynthetic sn-glycerol-3-phosphate dehydrogenase (EC 1.1.1.8) of Escherichia coli was potently inhibited by palmitoyl-CoA and other long chain acyl-CoA thioesters . The concentration dependence of this inhibition was not cooperative . Enzyme activity was inhibited 50% at 1 microM palmitoyl-CoA; thus, this inhibition occurred at concentrations below the critical micellar concentration of palmitoyl-CoA . Palmitoyl-CoA was a reversible, noncompetitive inhibitor with respect to both NADPH and dihydroxyacetone phosphate . Palmitoyl-CoA did not affect the quaternary structure of the enzyme . This inhibition could be prevented or reversed by the addition of phospholipid vesicles prepared from E . coli phospholipids . Palmitoyl-CoA did not alter the kinetics of inhibition by sn-glycerol 3-phosphate, which is a proven physiological regulator of this enzyme . Decanoyl-CoA, dodecanoyl-CoA, myristoyl-CoA, palmitoyl-(1,N6-etheno)CoA, stearoyl-CoA, and oleoyl-CoA inhibited sn-glycerol-3-phosphate dehydrogenase at concentrations below their critical micellar concentrations . Palmitate inhibited sn-glycerol-3-phosphate dehydrogenase activity 50% at 200 microM . Palmitoyl-carnitine, deoxycholate, taurocholate, and dodecyl sulfate were more potent inhibitors than Triton X-100, Tween-20, or Tween-80 . Palmitoyl-acyl carrier protein at concentrations up to 50 microM had no effect on sn-glycerol-3-phosphate dehydrogenase activity . The possible physiological role of long chain fatty acyl-CoA thioesters in the regulation of sn-glycerol 3-phosphate and phospholipid biosynthesis in E . coli is discussed. J Biol Chem, 1979 Feb 25, 254(4), 1013 - 5 Evidence for a direct role of tRNA in an amino acid transport system; Yoo SH et al.; The transport of phenylalanine by the general aromatic transport system in spheroplasts of Escherichia coli 9723 has been found to be stimulated by exogenous tRNA . Neither periodate-treated tRNA nor phenylalanine-charged tRNA stimulated, and the latter inhibited, phenylalanine uptake . Among preparations of specific tRNAs, tRNAPhe and tRNATyr were effective in stimulating the uptake of phenylalanine and tyrosine, respectively, and tRNAGlu and tRNAVal gave no detectable stimulation of phenylalanine or tyrosine transport . The preparation of tRNATyr was 10 times as active as unfractionated tRNA and gave as much as 167% stimulation of tyrosine transport . Correspondingly, the preparation of tRNAPhe was at least 3.5 times as active as the unfractionated tRNA and 2.5 times as active as the preparation of tRNATyr in stimulation of phenylalanine transport . Preliminary results in fractionation of the active component of tRNA for stimulating phenylalanine uptake show that the major activity resides in minor isoacceptor(s) tRNAPhe rather than the major component tRNAPhe, and the slight activity of preparations of tRNATyr is probably due to a contamination of the active tRNAPhe . Other preliminary results indicate that this type of stimulation occurs with uptake of other amino acids and their tRNA. Biochim Biophys Acta, 1979 Feb 20, 551(1), 157 - 68 Unmasking of an essential thiol during function of the membrane-bound enzyme II of the phosphenolpyruvate beta-glucoside phosphotransferase system of Escherichia coli; Haguenauer-Tsapis R et al.; beta-Glucoside transport by phosphoenolpyruvate-hexose phosphotransferase system in Escherichia coli is inactivated in vivo by thiol reagents . This inactivation is strongly enhanced by the presence of transported substrates . In a system reconstituted from soluble and membrane-bound components, only the particulate component, the membrane-bound enzyme IIbgl appeared as the target of N-ethylmaleimide inaction . The same feature was found in the case of methyl-alpha-D-glucoside uptake via enzyme IIglc . It is shown that the sensitizing effect of substrates is specific and not generalized, methyl-alpha-D-glucoside only sensitizes enzyme IIglc and p-nitrophenyl-beta-D-glucoside only sensitizes enzyme IIbgl towards N-ethylmaleimide inactivation . The inactivation of enzyme IIbgl by thiol reagents is also promoted in vivo by fluoride inhibition of phosphoenolpyruvate synthesis . In toluene-treated bacteria, the presence of phosphoenolpyruvate protects against inactivation by thiol reagents of p-nitrophenyl-beta-D-glucoside phosphorylation . Both results suggest that the inactivator resistent form of enzyme IIbgl is an energized form of the enzyme. Biochemistry, 1979 Feb 20, 18(4), 637 - 47 Photocross-linking analysis of the contact surface of tRNA Met in complexes with Escherichia coli