Escherichia coli

J Clin Chem Clin Biochem, 1977 Aug, 15(8), 433 - 8
{Metabolism and action of intracellular magnesium (author's transl)}; Gunther T; The activity of Mg2+-activated enzymes shows a bell shaped pMg dependency, with a pMg optimum at 3 . Intracellular Mg2+ concentrations, determined by different methods, are about 10(-3) mol/1 . Thus Mg2+-dependent enzymes are optimally activated, or nearly so, with Mg2+ . If the substrates of an enzyme form complexes with Mg2+ of different stabilities, and if free substrate and the substrate-Mg2+ complex react differently with the enzyme, the equilibrium will change with the concentration of Mg2+ . When the extracellular concentration of Mg2+ is increased, Mg2+ becomes bound practically exclusively to the cell membrane . In Mg2+ deficiency, the intracellular concentrations of Na+, K+, Ca2+ and cycl . AMP are changed, and the rates of synthesis of DNA, RNA and protein are decreased.

Zentralbl Bakteriol {Orig B}, 1977 Aug, 164(5-6), 492 - 7
{Detection of viruses in water of the Baltic Sea (author's transl)}; Steinman J; Virological examination of water of the Baltic Sea in the neighbourhood of a sewage outfall was done . By means of an apparatus for concentrating viruses in water, it was possible to detect enteroviruses in four of eleven samples, and in general in those moments, when conditions were fulfilled by a certain windway . The amount of viruses varied from 5 to 126 pfu in 10 liters, the ratio of virus to E . coli titer from 1:11 111 to 1:100 000 . Factors influencing the decrease of virus titer in seawater were briefly discussed.

Proc Natl Acad Sci U S A, 1977 Aug, 74(8), 3226 - 9
Nucleotide phosphotransferase of Escherichia coli: purification by affinity chromatography; Brunngraber EF et al.; Improved extraction and purification procedures permit the isolation from Escherichia coli W cells of much larger quantities and of more highly purified preparations of nucleotide phosphotransferase . Of various affinity resins tested for efficiency of purification, columns of agarose/5'-AMP (AGAMP), type 3, proved the best . In this way a 300- to 450-fold purification of the enzyme was achieved in a few steps . The enzyme, which, as reported before, transfers organically bound phosphate to the 2' or 3' hydroxyls of nucleosides and nucleotides, was tested in its behavior toward a series of ribonucleosidonucleotides, namely, CpC, ApA, CpA, and ApC . All were phosphate acceptors, but a detailed comparative study of adenosine and cytidine, 5'-AMP and 5'-CMP, and ApA and ApC revealed peculiar specificities in the relative distribution of the phosphorylated products.

Can J Biochem, 1977 Aug, 55(8), 911 - 5
Some biochemical effects of 4-deoxy-4-fluoro-D-glucose on Escherichia coli; Taylor NF et al.; The uptake of 4-deoxy-4-fluoro-D-glucose (4FG), without subsequent catabolism, by resting cells of Escherichia coli (ATCC 11775) is 0.06 mg/mg dry weight . In frozen-thawed cells of this organism, 4FG is a substrate for the phosphoenolpyruvate phosphotransferase system with a rate of phosphorylation twice that found for the isomeric 3-deoxy-3-fluoro-D-glucose . 4FG is not a carbon source for growth of this organism and it inhibits the extent of growth of cells in the presence of glucose . The inhibition of growth of E . coli K12 on lactose by 4FG is also observed and this is considered to be consistent with the fact that 4FG is an uncompetitive inhibitor of beta-galactosidase (EC 3.2.1.23) activity and that 4FG or 4-deoxy-4-fluoro-D-glucose-6 phosphate repress beta-galactosidase synthesis . These results support the view that catabolite repression may be produced by compounds which are not necessarily metabolised further than hexose-6-phosphates.

J Bacteriol, 1977 Aug, 131(2), 707 - 9
Role of cyclic adenosine 3',5'-monophosphate on cessation of respiration in ultraviolet-irradiated Escherichia coli; Swenson PA et al.; The addition of cyclic adenosine 3',5'-monophosphate (cAMP) to ultraviolet-irradiated Escherichia coli B/r cultures causes additional cells to cease respiring and to die . These effects of cAMP are greater on glucose-grown cells, where the effects of ultraviolet radiations alone are smaller and where the intracellular concentrations of cAMP are known to be lower.

Infect Immun, 1977 Aug, 17(2), 286 - 9
Protective capacity of antibodies against Escherichia coli and K antigens; Kaijser B et al.; Antibodies to Escherichia coli O and K antigens were raised in rabbits by repeated immunizations with whole, Formalin-killed and, later, liver bacteria . The serum antibody levels were determined with the ammonium sulfate precipitation technique after radioiodinating the antigens . The K antigens had to be conjugated to proteins before labeling . Such conjugations were performed using cyanogen bromide for the K1 antigen and bisdiazobenzidine for the K13 antigen . The protective capacities of the rabbit antisera were tested in intraperitoneally infected mice . The protective capacity of the antisera was expressed per ammonium sulfate precipitation titer . The results showed a significantly higher protective effect for the antibodies against the K1 and K13 antigens than for the antibodies against the O2 and O6 lipopolysaccharides.

Proc Natl Acad Sci U S A, 1977 Aug, 74(8), 3167 - 70
Sodium-stimulated glutamate uptake in membrane vesicles of Escherichia coli: the role of ion gradients; MacDonald RE et al.; Membrane vesicles prepared from Escherichia coli B/r grown on glutamate as a sole source of carbon and energy require sodium for glutamate accumulation when energized by D-lactate oxidation . Glutamate uptake can also be driven by a prearranged sodium gradient (out to in) in the absence of an energy source or a protonmotive force . Sodium ions are exchanged rapidly in respiring vesicles and the sodium gradient may be large enough under certain conditions to drive glutamate uptake after the protonmotive force is abolished with m-chlorocarbonylcyanide phenylhydrazone . Glutamate uptake due to a prearranged sodium gradient or lactate oxidation is inhibited by monensin but not by nigericin . Transport does not occur in response to valinomycin-induced membrane potential . We interpret these results to indicate that glutamate transport is obligately coupled to sodium transport and can only occur when there is a net flux of sodium ions . This flux is driven by a chemical gradient of sodium that is created by the protonmotive force generated by respiration.

Nucleic Acids Res, 1977 Aug, 4(8), 2931 - 8
Enzymatic synthesis of Q nucleoside containing mannose in the anticodon of tRNA: isolation of a novel mannosyltransferase from a cell-free extract of rat liver; Okada N et al.; The Q nucleosides isolated from rabbit liver tRNA are known to have sugars (mannose or galactose) linked to their cyclopentene diol moiety . A Q nucleoside containing mannose (manQ) was synthesized by a cell-free system from rat liver, using purified E . coli tRNAAsp as an acceptor and GDP-mannose as a donor molecule . The novel mannosyltransferase catalyzing this reaction was purified from a particulate-free soluble enzyme fraction and found to be strictly specific for tRNAAsp . These results, together with the anomeric configuration of mannose in Q nucleoside, indicate that no lipid intermediate is involved in the biosynthesis of Q nucleoside.

Am J Physiol, 1977 Aug, 233(2), E71 - 9
Glucose utilization and role of blood in endotoxin shock; Hinshaw LB et al.; The present study was conducted to explore influences modifying glucose uptake in canine blood administered LD100 E . coli endotoxin . Particular emphasis was given to assay the role of the white blood cell (WBC) in glucose utilization . Significant increases in glucose uptake and lactic acid production, attributed to increased activity of the WBC, were observed 1-3 h after endotoxin was added to blood in vitro . Although a net increase in glucose utilization was noted, endotoxin simultaneously exerted adverse effects by depressing glucose uptake below predicted values (Q10 = 2.12 with LD100 endotoxin vs . 2.78 in saline controls) and increasing WBC mortality rate . Blood from dogs pretreated with sublethal doses of endotoxin in vivo utilized glucose at an accelerated rate when subjected to endotoxin in vitro . Excess glucose was consumed because of elevated numbers of white blood cells although additional glucose requirements after endotoxin were independent of temperature between the ranges of 34-41 degrees C . All animals pretreated with daily sublethal injections of endotoxin for 3 days survived superlethal doses of endotoxin.

Science, 1977 Jul 29, 197(4302), 452 - 5
Dihydrofolate reductase: x-ray structure of the binary complex with methotrexate; Matthews DA et al.; A central eight-stranded beta-pleated sheet is the main feature of the polypeptide backbone folding in dihydrofolate reductase . The innermost four strands and two bridging helices are geometrically similar to but are connected in a different way from those in the dinucleotide binding domains found in nicotinamide-adenine dinucleotide-linked dehydrogenases . Methotrexate is bound in a 15-angstrom-deep cavity with the pteridine ring buried in a primarily hydrophobic pocket, although a strong interaction occurs between the side chain of aspartic acid 27 and N(1), N(8), and the 2-amino group of methotrexate.

Biochemistry, 1977 Jul 26, 16(15), 3465 - 9
Effect of proteolysis of transcriptional fidelity of reconstituted chromatin; Gadski RA et al.; The effect of proteolysis on the transcriptional properties of reconstituted rat liver chromatin was studied . Within the sensitivity of currently available methods, proteolysis of chromosomal proteins by chromatin-bound protease during chromatin reconstitution has no apparent effect on: (1) number of initiation sites, (2) proportion of reiterating and unique sequences of DNA transcribed, (3) size of the RNA transcribed, and (4) transcription of DNA sequences complementary to poly(A) containing messenger RNA.

Biochemistry, 1977 Jul 26, 16(15), 3334 - 42
Subunit topography of RNA polymerase from Escherichia coli . A cross-linking study with bifunctional reagents; Hillel Z et al.; The quaternary structures of Escherichia coli DNA-dependent RNA polymerase holenzyme (alpha 2 beta beta' sigma) and core enzyme (alpha 2 beta beta') have been investigated by chemical cross-linking with a cleavable bifunctional reagent, methyl 4-mercaptobutyrimidate, and noncleavable reagents, dimethyl suberimidate and N,N'-(1,4-phenylene)bismaleimide . A model of the subunit organization deduced from cross-linked subunit neighbors identified by dodecyl sulfate-polyacrylamide gel electrophoresis indicates that the large beta and beta' subunits constitute the backbone of both core and holoenzyme, while sigma and two alpha subunits interact with this structure along the contact domain of beta and beta' subunits . In holoenzyme, sigma subunit is in the vicinity of at least one alpha subunit . The two alpha subunits are close to each other in holoenzyme, core enzyme, and the isolated alpha 2 beta complex . Cross-linking of the "premature" core and holoenzyme intermediates in the in vitro reconstitution of active enzyme from isolated subunits suggests that these species are composed of subunit complexes of molecular weight lower than that of native core and holoenzyme, respectively . The structural information obtained for RNA polymerase and its subcomplexes has important implications for the enzyme-promoter recognition as well as the mechanism of subunit assembly of the enzyme.

J Biol Chem, 1977 Jul 25, 252(14), 5019 - 31
Human beta-globin messenger RNA . I . Nucleotide sequences derived from complementary RNA; Marotta CA et al.; Sequence analysis studies were carried out on human beta-globin mRNA (beta-mRNA) prepared from alpha-thalassemic, sickle cell, and Hb A reticulocytes . Highly purified beta-mRNA served as substrate for the preparation of cDNA by RNA-dependent DNA polymerase . The cDNA was transcribed by Escherichia coli RNA polymerase and the resulting cRNA was analyzed . Over 300 nucleotides were assigned to the beta-mRNA coding region and 37 nucleotides were assigned to the 3'-terminal noncoding region . The normal termination codon is UAA which is separated by 28 nucleotides from an out of phase UAA triplet . The origin of each of the abnormally long beta-globin variants Tak and Cranston is consistent with reduplication of dinucleotides prior to the normal termination codon, and both globin variants can terminate at the out of phase UAA.

J Biol Chem, 1977 Jul 25, 252(14), 4786 - 9
Novel properties of Escherichia coli exonuclease III; Roychoudhury R et al.; The specificity of hydrolysis of polynucleotide termini by Escherichia coli exonuclease III was studied with the use of oligothymidylate annealed to polydeoxyadenylate . The size of the products after 3' leads to 5'-hydrolysis of 5'-labeled substrate is temperature-dependent . At 25 degrees the enzyme can hydrolyze a polynucleotide chain up to the last 5'-terminal dinucleotide . A gradation of higher 5'-terminal oligonucleotides of defined chain lengths is produced after limit digestion by the enzyme when the temperature is raised between 25 degrees to 60 degrees . When the oligothymidylate was labeled at the 3'-ends with ribonucleotides, it was observed that exonuclease III can cleave a single or two consecutive ribonucleotides regardless of whether the ribonucleotides are base-paired or mismatched.

J Biol Chem, 1977 Jul 25, 252(14), 4749 - 51
Studies of the lipid phase transitions of Escherichia coli by high sensitivity differential scanning calorimetry; Jackson MB et al.; High sensitivity adiabatic differential scanning calorimetry was performed on lipids, membrane vesicles, and whole cells of Escherichia coli enriched in particular unsaturated fatty acids by genetic means . Information concerning the shape of the transition is discussed . Transitions with an asymmetric shape reminiscient of a second order transition were observed . Comparison between the lipid transition observed in whole cells, membrane vesicles, and extracted lipids enriched in elaidate reveal some basic similarities . Studies of synthetic lipids were undertaken in an attempt to interpret the shapes of these transitions as a function of the lipid components of the membrane.

J Biol Chem, 1977 Jul 25, 252(14), 4790 - 5
Purification and properties of tRNA(adenine-1)-methyltransferase from rat liver; Glick JM et al.; An S-adenosylmethionine-dependent tRNA(adenine-1)-methyltransferase has been purified 8,000-fold from rat liver . This preparation gives a single band on polyacrylamide gel electrophoresis and is stable in long term storage . The enzyme has a molecular weight of approximately 95,000 . The single methylating capacity of this adenine-1 methyltransferase, using Escherichia coli tRNA2Glu, is methylation of the invariant adenine in the GTpsiC loop . The methylation reaction is dependent on added cation with 20 to 40 mM putrescine being most effective . The Km for S-adenosylmethionine was found to be 0.3 micron, while the Ki for the product inhibitor S-adenosylhomocysteine was 0.85 micron . The Km for tRNAMetf is 12 nM while that for tRNAGlu2 is 33 nM.

Schweiz Med Wochenschr, 1977 Jul 23, 107(29), 1028 - 34
{Activation of the complement system in different forms of glomerulonephritis}; Wegmuller E et al.; The complement system may be activated by two pathways, the classical and the alternate . To evaluate their respective participation in different forms of glomerulonephritis, the plasma values of C3, C4, C3PA, C1q and properdin were determined in 70 patients . In systemic lupus erythematosus (LED), acute poststreptococcal glomerulonephritis (AGN) and septicemia the classical pathway appears to be mainly involved, whereas the amplification loop and the alternate pathway seem to be of secondary importance . By contrast, in membranoproliferative glomerulonephritis (MPGN) the alternate pathway plays a major role . However, the present data suggest that activation of the classical pathway may often be involved as well . In minimal change glomerulonephritis no signs indicating involvement of the complement system were apparent . Follow-up observation demonstrated a correlation between decreases in plasma complement concentrations and the clinical severity of the primary disease in LED, AGN and septicemia, but not in MPGN.

Biochim Biophys Acta, 1977 Jul 22, 493(1), 210 - 5
Purification of protein A, an outer membrane component missing in Escherichia coli K-12 ompA mutants; Chai TJ et al.; Outer membrane materials prepared from an Escherichia coli ompA (tolG) strain do not contain one of the major outer membrane proteins found in ompA+ strains . This protein has been purified in high yield from detergent-solubilized cell envelope material prepared from an ompA+ strain by preparative electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate . The purified protein is homogeneous in three electrophoretic systems, contains 2 mol of reducing sugar/mol of peptide and has alanine as the N-terminal amino acid . The amino acid composition is nearly identical to outer membrane protein II or B purified by others from incompletely solubilized cell envelope material . Thus, the fraction of outer membrane protein II or B that is difficult to solubilize is identical with the more readily solubilized fraction.

Mol Gen Genet, 1977 Jul 20, 154(2), 205 - 11
Transposition of TnA does not generate deletions; Bennett PM et al.; We have examined the incidence of loss of the TnA unit, Tn801, from RP1 under conditions where transposition of Tn801 to another replicon . R388, was readily detected . We found that the frequency of transposition of Tn801 from RP1 to R388 exceeded, by at least a factor of one hundred, the frequency at which it was deleted from RP1 . We conclude that, in general, transposition of Tn801 does not generate derivatives of the donor plasmid which specifically lack Tn801 . The relevance of these findings to the mechanism of transposition is discussed.

Mol Gen Genet, 1977 Jul 20, 154(2), 185 - 9
Arsenate-resistant alkaline phosphatase-constitutive mutants of Escherichia coli; Yagil E et al.; When arsenate-resistant mutants are selected approximately 50 per cent of them are also consitutive for the synthesis of alkaline phosphatase and the Pi-binding protein . Some of these mutants are linked to ilv (phoS- or phoT-), other are linked to proC (phoR-) . One of the mutant strains linked to ilv lost the Pi-binding protein (the phoS gene product) . Resistance to arsenate, constitutivty for alkaline phosphatase synthesis and loss of the Pi-binding protein occurred pleiotropically by the same phoS- mutation.

Mol Gen Genet, 1977 Jul 20, 154(2), 175 - 80
Genetics of ribosomal protein methylation in Escherichia coli . II . A mutant lacking a new type of methylated amino acid, N5-methylglutamine, in protein L3; Lhoest J et al.; The ribosomes of an Escherichia coli mutant, designated prm-2, can be methylated in vitro by an enzymatic fraction from wild-type . This enzyme is inactive on the ribosomes from another mutant, prm-1, is reported previously to be methyl group-deficient in protein L11 . In vitro methylation of prm-2 ribosomes resulted in the incorporation of about one methyl group per molecule of protein L3 . After acid hydrolysis, all the methyl groups were found in a very basic compound which was identified as methylamine . This compound could have been generated by acid hydrolysis of N-methylated amide-groups from glutamine or asparagine . Therefore, chemically-synthesized N4-methyl-asparagine and N5-methylglutamine were chromatographed together with an enzymatic hydrolysate of methylated prm-2 proteins . In all the chromatogrphic systems studied the methylated amino acid was found in the same position as N5'-methylglutamine . These results indicate that mutant prm-2 lacks one residue of N5-methylglutamine present in ribosomal protein L3 of wild type E . coli.

Mol Gen Genet, 1977 Jul 20, 154(2), 167 - 73
Genetics of ribosomal protein methylation in Escherichia coli . I . A mutant deficient in methylation of protein L11; Colson C; Several thousand mutagenized clones of Escherichia coli were screened for methyl group incorporation into protein in crude extracts, in order to isolate mutants lacking the full complement of methyl groups in ribosomal proteins . One mutant isolated by this method and designated prm-1 incorporated 6-7 methyl groups per ribosome upon incubation of its ribosomes with a partially purified enzyme preparation from E . coli wild-type . The methyl groups were located exclusively in the 50S particle and for the most part (85%) in protein L11 . Three methylated amino acids were detected: epsilon-N-trimethyllysine, epsilon-N-monomethyllysine, and an uncharacterized amino acid . These accounted respectively for 4.6, 1.3 and 0.9 methyl groups per ribosome . These results indicate that protein L11 in wild-type contains a stoichiometric amount of these methylated amino acids which are absent in mutant prm-1 . Since this mutant is fully viable, its methylation deficiency does not result in a major defect in ribosome assembly or functioning.

Mol Gen Genet, 1977 Jul 20, 154(2), 113 - 21
Escherichia coli plasmid pBR313 insertion into plant protoplasts and into their nuclei; Lurquin PF et al.; Cowpea mesophyll protoplasts were shown to bind irreversibly up to 3% input radioactive pBR313 plasmid DNA after 15 min of contact . Maximum uptake occurred in the presence of 5 mM ZnSO4 and 5 microgram/ml poly-L-ornithine . Under these conditions about one half of the TCA precipitable radioactivity was associated with the nuclear fraction and behaved as linear plasmid molecules . These could not be chased from the protoplasts upon further incubation with unlabeled plasmid DNA . The presence of donor DNA within the nuclear fraction is most probably not due to an artifactual redistribution of adsorbed plasmid DNA . Prolonged incubation periods resulted in extensive degradation of plasmid in the incubation medium but little degradation occurred in the protoplasts . The donor DNA was not covalently associated with the protoplast nuclear DNA.

Biochim Biophys Acta, 1977 Jul 15, 477(2), 102 - 11
Inhibition of amino acyl tRNA synthetase activity by copper complexes of two metal binding ligands . N-Methyl isatin beta-thiosemicarbazone and 8-hydroxyquinoline; Rohde W et al.; Copper complexes of N-methyl isatin beta-thiosemicarbazone, 1-formyl isoquinoline thiosemicarbazone and thiosemicarbazide inhibit amino acyl tRNA synthetase activity . Copper complexes of 8-hydroxyquinoline and 8-mercaptoquinoline also inhibit . The 1 : 1 ligand-metal complex is significantly more active than the 2 : 1 complex . The free ligand alone and copper sulfate alone have little, if any, effect . These complexes have no effect on the ATP-PPi exchange reaction and do not cause deacylation of amino acyl tRNAs . This indicates that the process inhibited by these complexes is the amino acylation reaction . This is the first report that these copper binding ligands can inhibit enzymatic processes which involve nucleic acids but which are not viral, bacterial or mammalian cell polymerases.

Eur J Biochem, 1977 Jul 15, 77(2), 217 - 22
Mapping of 23-S rRNA at the ribosomal peptidyl-transferase center by photo-affinity labeling; Sonenberg N et al.; Photo-sensitive peptidyl-tRNA's were used to scan the environment of the peptidyl-transferase center of the ribosome . The specificity of the previously described labeling in the 18-S fragment of 23-S rRNA by Boc-Phe(N3)-Phe-tRNA (4-azido-N-t-butoxycarbonyl-phenylalanyl-phenylalanyl-tRNA) was demonstrated by the ability of the covalently anchored molecule to serve as donor substrate in peptide bond formation . Labeling patterns were also obtained with Boc-Phe(N3)-Phe-Phe-tRNA bound at the acceptor site and with Boc-Phe(N3)-(Gly)n-Phe-tRNA (n = 2,4) . The results indicate that subsequences within the 18-S fragment of 23-S rRNA are located close to the acceptor site as well as along the path where the peptide moiety adheres to the ribosome . Identification of the labeled sequences is expected to shed light on the spatial arrangement as well as functional role of rRNA in the peptidyl transferase center.

Biochim Biophys Acta, 1977 Jul 15, 477(2), 97 - 101
A comparison of the differential DNA melting profiles with the CsCl density profiles of DNA from Escherichia coli, cow, mouse, rat and chicken; Mayfield JE; Moderate resolution thermal denaturation profiles are presented for the purified DNAs from Escherichia coli, cow, mouse, rat and chicken . All show multiple thermal transitions indicative of large blocks of DNA with very similar base composition . The eucaryotes all have much more of this kind of DNA than does E . coli . The satellite DNAs of cow and mouse are clearly visible and it is likely that the other transitions represent additional families of repeated DNA.

Biochim Biophys Acta, 1977 Jul 15, 477(2), 144 - 50
Inhibition of DNA polymerase-alpha and -beta of calf thymus by 1-beta-D-arabinofuranosylcytosine-5'-triphosphate; Yoshida S et al.; 1-beta-D-Arabinofuranosylcytosine 5'-triphosphate (araCTP), an active form of a inhibitor of DNA replication, 1-beta-D-arabinofuranosylcytosine (araC) was tested for its inhibitory action on the DNA polymerase-alpha and -beta (EC 2.7.7.7) purified from calf thymus . The reaction of DNA polymerase-alpha was shown to be more sensitive to the inhibition by araCTP than that of DNA polymerase-beta . The mode of the inhibition by araCTP was competitive to dCTP in the reaction catalysed by either DNA polymerase-alpha or -beta . The Ki value of DNA polymerase-beta for araCTP was 32 micron; eight times higher than that of DNA polymerase-alpha (4 micron) for this inhibition.

Science, 1977 Jul 15, 197(4300), 261 - 3
Mechanism of carbon isotope fractionation associated with lipid synthesis; DeNiro MJ et al.; The low carbon-13/carbon-12 ratio of lipids is shown to result from isotopic fractionation during the oxidation of pyruvate to acetyl coenzyme A . In vitro analysis of the kinetic isotope effects of this reaction indicates that there will be a large, temperature-dependent difference in the carbon-13/carbon-12 ratio between the methyl and carbonyl carbon atoms of acetyl coenzyme A and between those carbon atoms of lipid components which derive from them.

Biochemistry, 1977 Jul 12, 16(14), 3133 - 6
Template activity of calf thymus DNA modified by a dihydrodiol epoxide derivative of benzo{a}pyrene; Leffler S et al.; The purpose of the present study was to determine the effects of covalent binding to DNA of a reactive derivative of benzo{a}pyrene on template activity during in vitro transcription with RNA polymerase . Calf thymus deoxyribonucleic acid, modified by reaction with (+/-)-7beta,8alpha-dihydroxy-9alpha, 10alpha-epoxy-7,8,9,10-tetrahydrobenzo{a}pyrene, was transcribed with Escherichia coli DNA-dependent RNA polymerase . With increasing levels of modification, there was a progressive inhibition of transcription . The inhibition was much greater under conditions where continuous reinitiation of transcription occurred than under conditions where only one RNA chain was synthesized per initiation site . This suggested that the modified sites block the movement of polymerase along the template and prevent recycling of the enzyme . Consistent with this interpretation were analyses of RNA transcripts on sucrose density gradients which showed a progressive decrease in average RNA chain length as the extent of template modification increased . In contrast to the inhibitory effect on chain elongation, evidence was obtained that the modified DNA had an increase in the number of initiation sites for transcription . These results are consistent with separate physical studies indicating that modification of DNA by this benzo{a}pyrene derivative can induce small localized regions of denaturation.

Biochemistry, 1977 Jul 12, 16(14), 3256 - 61
Preparation and properties of a new DNase from Aspergillus oryzae; Rushizky GW et al.; A DNase present in commercial preparations of Aspergillus oryzae alpha-amylase was purified 1550-fold in 25% yield by acetone precipitation and by chromatography on diethylaminoethyl- and carboxymethylcellulose . The enzyme was isolated free of contaminating RNases and DNases . The molecular weight of the enzyme determined by gel filtration on Sephadex G-100 was 48 000, while a molecular weight of 58 000 was determined for the single band observed upon polyacrylamide gel electrophoresis in sodium dodecyl sulfate . The isoelectric point of the DNase is 9.2 . The enzyme hydrolyzed only DNA with a pH optimum of 8.2 and was activated by Co2+, and to a lesser extent by Mg2+ and Mn2+ . Native DNA was a better substrate than heat-denatured DNA . Enzymatic digests of calf thymus and E . coli DNA yielded oligomers of chain lengths ranging from 10 to 200, with mono- and small oligonucleotides (chain length less than 5) detected only when large (100 mg) amounts of DNA were fractionated by column chromatography on diethylaminoethyl-Sephadex A-25 in 7 M urea . The digestion products contained 5'-terminal phosphate groups and mostly adenosine at the 3' and guanosine and adenosine at the 5' ends.

J Biol Chem, 1977 Jul 10, 252(13), 4435 - 7
Release factor binding to ribosome requires an intact 16 S rRNA 3' terminus; Caskey CT et al.; Cloacin DF12 cleavage of Escherichia coli f{3H}MettRNA-AUG-ribosome complexes affects this substrate for in vitro peptide chain termination . Codon-directed release factors' (RF) 1 and 2 release of f{3H}methionine is inhibited by cloacin . Since cloacin inhibits RF1 and -2 binding to ribosomes but not RF-directed f{3H}methionine release from f{3H}met-tRNA-AUG-ribosome complexes when reactions contain 20% ethanol, we conclude that cloacin DF 13 inhibits formation of the termination codon recognition complex . Thus, cleavage of the 3'-OH 49-nucleotide sequence of the 16 S rRNA perturbs the codon-directed binding of RF to ribosomes.

Chromosoma, 1977 Jul 8, 62(3), 199 - 215
Interactions stabilizing DNA tertiary structure in the Escherichia coli chromosome investigated with ionizing radiation; Lydersen BK et al.; The structure of the bacterial chromosome was investigated after introducing breaks in the DNA with gamma irradiation . It is demonstrated that irradiation of the chromosome in the cell prior to isolation results in partial unfolding of the isolated condensed DNA, while irradiation of the chromosome after it is released from the cell has no demonstrable effect on DNA folding . The results indicate that RNA/DNA interactions which stabilize DNA folds are unstable when breaks are introduced in the DNA prior to isolation of the chromosome . It is suggested that the supercoiled state of the DNA is required for the initial stabilization of some of the critical RNA/DNA interaction in the isolated nucleoid . However, some of these interactions are not affected by irradiation of the cells . Remnant supercoiling in partially relaxed chromosomes containing a limited number of DNA breaks has the same superhelical density as the unirradiated chromosome . This suggests that restraints on rotation of the packaged DNA are formed prior to the physical unwinding which occurs at the sites of the radiation induced DNA breads . - Analysis of the in vitro irradiated chromosomes shows that there are 100 +/- 30 domains of supercoiling per genome equivalent of DNA . The introduction of up to 50 double-strand breaks per nucleoid does not influence rotor speed effects of the sedimentation coefficient of the chromosome.

Biochim Biophys Acta, 1977 Jul 8, 483(1), 24 - 34
Concerted inhibition of NADP+-specific isocitrate dehydrogenase by oxalacetate and glyoxylate . I . Oxalomalate formation and stability, and nature of the enzyme inhibition; Johanson RA et al.; Oxalacetate and glyoxylate are each weak inhibitors of NADP+-specific isocitrate dehydrogenase (threo-DS-isocitrate:NADP+ oxidoreductase (decarboxylating), EC 1.1.1.42)9 Together, however, they act in a concerted manner and strongly inhibit the enzyme . The rates of formation and dissociation of the enzyme inhibitor complex, and the rate of formation and the stability of the aldol condensation product of oxalacetate and glyoxylate, oxalomalate, were examined . The data obtained do not support the often suggested possibility that oxalomalate, per se, formed non-enzymatically in isocitrate dehydrogenase assay mixtures containing oxalacetate and glyoxylate, is responsible for the observed inhibition of the enzyme . Rather, the data presented in this communication suggest that oxalacetate binds to the enzyme first, and that the subsequent binding of glyoxylate leads to the formation of a catalytically inactive enzyme-inhibitor complex.

Mol Gen Genet, 1977 Jul 7, 154(1), 83 - 6
Ribosomal abnormality in recA mutants of Escherichia coli; Powell KA et al.; The tif-1 mutation has been shown to affect protein synthesis in vitro by increasing translational ambiguity (Ephrati-Elizur, Luther-Davies and Hayes, 1976) . It is demonstrated here that some recA mutations confer similar abnormality . By comparing suitable combinations of ribosomes and soluble proteins from recA+ and recA cells the defect is shown to be associated with ribosomes . The recA mutation, which suppresses most phenotype characteristics of the tif-1 mutation (Castellazzi, George and Buttin, 1972(b)) does not suppress the ribosomal abnormality . Since the closely linked tif-1 and recA mutations lead to the expression of a common property they may be in the same gene.

Biochim Biophys Acta, 1977 Jul 7, 461(1), 84 - 100
Role of quinones in electron transport to oxygen and nitrate in Escherichia coli . Studies with a ubiA- menA- double quinone mutant; Wallace BJ et al.; A ubiA- menA- double quinone mutant of Escherichia coli K12 was constructed together with other isogenic strains lacking either ubiquinone or menaquinone . These strains were used to study the role of quinones in electron transport to oxygen and nitrate . Each of the four oxidases examined (NADH, D-lactate, alpha-glycerophosphate and succinate) required a quinone for activity . Ubiquinone was active in each oxidase system while menaquinone gave full activity in alpha-glycerophosphate oxidase, partial activity in D-lactate oxidase but was inactive in NADH and succinate oxidation . The aerobic growth rates, growth yields and products of glucose metabolism of the quinone-deficient strains were also examined . The growth rate and growth yield of the ubi+menA- strain was the same as the wild-type strain, whereas the ubiA-men+ strain grew more slowly on glucose, had a lower growth yield (30% of wild type) and accumulated relatively large quantities of acetate and lactate . The growth of the ubiA-menA- strain was even more severely affected than that of the ubiA-men+ strain . Electron transport from formate, D-lactate, alpha-glycerophosphate and NADH to nitrate was also highly dependent on the presence of a quinone . Either ubiquinone or menaquinone was active in electron transport from formate and the activity of the quinones in electron transport from the other substrates was the same as for the oxidase systems . In contrast, quinones were not obligatory carriers in the anaerobic formate hydrogenlyase system . It is concluded that the quinones serve to link the various dehydrogenases with the terminal electron transport systems to oxygen and nitrate and that the dehydrogenases possess a degree of selectivity with respect to the quinone acceptors.

Biochim Biophys Acta, 1977 Jul 7, 461(1), 75 - 83
Aerobic respiration in mutants of Escherichia coli accumulating quinone analogues of ubiquinone; Wallace BJ et al.; The ability of three naturally occurring analogues of ubiquinone to function in aerobic respiration in Escherichia coli has been studied . The compounds, which differ from ubiquinone in terms of the substituents on the quinone ring, accumulate in the cytoplasmic membranes of ubiE-, ubiF- and ubiG- mutants . One of the analogues (2-octaprenyl-3-methyl-6-methoxy-1,4-benzoquinone, NMQ), which lacks the 5-methoxyl group of the benzoquinone ring of ubiquinone promoted the oxidation of NADH, D-lactate and alpha-glycerophosphate but not succinate . Electron transport supported by MMQ was found to be coupled to phosphorylation . In contrast, 2-octaprenyl-6-methoxy-1,4-benzoquinone, which lacks both the 3-methyl and 5-methoxyl groups of ubiquinone, and 2-octaprenyl-3-methyl-5-hydroxy-6-methoxy-1,4-benzoquinone, in which the 5-methoxyl group of ubiquinone is replaced by an hydroxyl group, were virtually inactive in the oxidases tested . The ability of MMQ to function in respiration in isolated membranes is consistent with the findings that the growth rate and yield of a ubiF- strain, unlike other ubi- strains, were only slightly lower than those of a ubiF+ strain . The fact that MMQ is active in some but not all oxidases provides further support for the concept that the quinones link the individual dehydrogenases to the respiratory chain and that each dehydrogenase has specific structural requirements for quinone acceptors.

Mol Cell Biochem, 1977 Jul 5, 16(2), 71 - 7
Polyamines and protein synthesis: studies in various polyamine-requiring mutants of Escherichia coli; Goldemberg SH et al.; Different Escherichia coli mutants auxotrophic for polyamines were studied in order to investigate the relationships among polypeptide synthesis in cell-free systems, ribosomal distribution profiles and endogenous polyamine pools . The in vitro protein synthetic activity and the polyribosomal content were reduced in extracts from putrescine-starved cells of the double mutans MA 255 and MA 261, but not in the arginine-conditional auxotroph DK 6 . Putrescine addition to the cultures of all these strains previously starved for polyamines, provoked a shift towards monomers in the equilibrium involving ribosomal particles . Concomitant changes in the intracellular levels of polyamines were observed: putrescine and spermidine increased markedly, and cadaverine disappeared.

Biochim Biophys Acta, 1977 Jul 5, 477(1), 84 - 8
Isoleucyl-tRNA synthetase inactivation and the extent of aminoacylation of tRNAIle from Escherichia coli; Marashi F et al.; A difference in isoleucine acceptance between normal and sulfur-deficient tRNA from Escherichia coli C6 (rel-, met-, cys-) was eliminated when more isoleucyl-tRNA synthetase was added at the reaction plateau . Enzymatic deacylation was similar for both tRNAs . These results suggest that enzyme inactivation caused a premature reaction plateau which was not predicted by the rates of acylation and deacylation.

Biochim Biophys Acta, 1977 Jul 5, 477(1), 20 - 7
Manganese(II) as a paramagnetic probe of the tertiary structure of transfer RNA; Chao YY et al.; The effect of manganese on both the low field (10--15 ppm) and the high field (o--3 ppm) NMR spectra of unfractionated tRNA and yeast tRNAPhe has been investigated . Trace amounts of Mn2+ cause selective broadening of resonances which are assigned to specific tertiary interactions . The order in which resonances broaden is the same as the order in which they are stabilized by the addition of magnesium, namely s4U8 - A14, U33 and A58 - T54 . From this we conclude that three of the strong binding sites probably are the same for both Mn2+ and Mg2+, and that these sites are located close to the tertiary interactions which are stabilized by the strongly bound metals . The broadening data, taken in conjunction with published X-ray data on yeast tRNAPhe, permit us to suggest some plausible locations for the strong binding sites.

Mol Cell Biochem, 1977 Jul 5, 16(2), 135 - 9
Influence of cyclic 3',5'-adenosine monophosphate on uracil uptake by rifampicin treated Escherichia coli cells; Judewicz ND et al.; Incubation of cells from a wild type strain of E . coli with 0.3 mg/ml rifampicin for 15 minutes lead to a complete inhibition of RNA synthesis measured as the uracil incorporation into the trichloroacetic acid insoluble fraction . In these rifampicin-treated cells {14C}uracil incorporation tended to decrease during a further incubation at 37 degrees . Addition of cyclic AMP increased the inactivation of the system responsible for {14C}uracil uptake . The cyclic nucleotide effect seems to be specific since ATP or 5'AMP did not increase such inactivation.

Biochim Biophys Acta, 1977 Jul 5, 477(1), 1 - 9
Involvement of NAD in induced synthesis of colicin E1; Nakazawa A et al.; Escherichia coli K-12 nadC13 (ColE1) was starved for nicotinic acid and cellular NAD levels decreased to less than 10% of the normal . Under these conditions, induction of colicin E1 synthesis decreased to about 1% of the normal value, while 30% of the total protein synthesis remained intact . Addition of nicotinic acid reversed both the ability of the colicin induction and cellular NAD level . Induced replication of colicin E1 DNA was greatly reduced in the NAD-deprived cells.

Biochim Biophys Acta, 1977 Jul 5, 477(1), 28 - 36
The effect of phenethyl alcohol on in vitro DNA synthesis in Escherichia coli; Kaneko M et al.; The effect of phenethyl alcohol on DNA synthesis was examined using several in vitro systems of Escherichia coli H560; i.e., ether-treated cells, membrane fractions and folded chromosomes fortified with DNA polymerase . In all systems, the incorporation of deoxyribonucleotides was much reduced for the phenethyl alcohol-treated cells compared with the non-treated cells . The total activity of DNA polymerases in polA1 cells (mostly DNA polymerase II) was not impaired for the phenethyl alcohol-treated cells and the reduction of the rate of DNA synthesis in vitro was ascribed to the reduction of the chromosomal template activity which was related to trypsin sensitive protein components . The analysis of chromosomes from the phenethyl alcohol-treated cells revealed the remarkable reduction of a protein component of molecular weight approx . 58 000 in contrast with a protein component of molecular weight approx . 30 000.

Res Vet Sci, 1977 Jul, 23(1), 84 - 6
In vitro stimulation of ovine lymphocytes by various mitogens; Burrells C et al.; A technique for the separation and in vitro culture of ovine lymphocytes is described and applied to the study of their blastogenic responses to various mitogens . Lymphocytes were separated on a Ficoll-Triosil gradient, resuspended in RPMI 1640 medium supplemented with 10 per cent serum to a concentration of 1 x 10(6) cells/ml and cultured in 200 microliter volumes in microculture plates in the presence of mitogens for varying lengths of time . A total culture period of 66 h was found to be satisfactory with 1 muCi of {3H}-thymidine being added to each well 18 h before termination of culture . Optimal blastogenic stimulation of the lymphocytes occurred with phytohaemagglutinin at 2-5 microliter/ml, pokeweed mitogen at 10 microliter/ml, concanavalin A at 6-25 microgram/ml and lipopolysaccharide of Escherichia coli (LPS) at 6-25 microgram/ml . Proliferation due to stimulation by LPS appeared to be of a lower order than that achieved with the other mitogens tested.

Biochem J, 1977 Jul 1, 165(1), 121 - 6
Affinity chromatography and inhibition of chorismate mutase-prephenate dehydrogenase by derivatives of phenylalanine and tyrosine; Smith GD et al.; Several derivatives of phenylalanine and tyrosine were prepared and tested for inhibition of chorismate mutase-prephenate dehydrogenase (EC 1.3.1.12) from Escherichia coli K12 (strain JP 232) . The best inhibitors were N-toluene-p-sulphonyl-L-phenylalanine, N-benzenesulphonyl-L-phenylalanine and N-benzloxycarbonyl-L-phenylalanine . Consequently two compounds, N-toluene-sulphonyl-L-p-aminophenylalanine and N-p-aminobenzenesulphonyl-L-phenylalanine, were synthesized for coupling to CNBr-activated Sepharose-4B . The N-toluene-p-sulphonyl-L-p-aminophenylalanine-Sepharose-4B conjugate was shown to bind the enzyme very strongly at pH 7.5 . The enzyme was not eluted by various eluents, including 1 M-NaCl, but could be quantitatively recovered by washing with buffer of pH9 . Elution was more effective in the presence of 10 mM-1-adamantaneacetic acid, a competitive inhibitor of the enzyme . This affinity-chromatography procedure results in a high degree of purification of the enzyme and can be used to prepare the enzyme in a one-step procedure from the bacterial crude extract . Such a procedure may therefore prove useful in studying this enzyme in a state that closely resembles that in vivo.

Clin Chem, 1977 Jul, 23(7), 1346 - 7
Heparin pretreatment suppresses norepinephrine concentrations in dogs in endotoxic shock; Devereux DF et al.; Mongrel dogs were treated intravenously with either 1000 units of beef-lung heparin per kilogram of body weight or with isotonic saline, before intravenous administration of E . coli endotoxin . We found significant differences in circulating norepinephrine concentrations between a heparin-pretreatment group (1.89 +/- 0.39 microgram/liter) and the control group (9.83 +/- 4.64 microgram/liter), but none with respect to epinephrine . Systolic blood pressures at 360 min were also significantly (P less than 0.05) different, 148 +/- 6 mmHg as compared with 118 +/- 13.4 mmHg . Evidently heparin pretreatment can decrease circulating norepinephrine concentrations in the endotoxic state and changes in circulating catecholamine concentrations can affect physiological variables.

Exp Pathol (Jena), 1977 Jul-Aug, 14(1-2), 33 - 9
Semiquantitative histological examinations of the kidneys of rabbits (64-day, 100-day and 212-day series) with experimentally induced pyelonephritis; Sorger K et al.; The kidneys of each 8 rabbits of a 64-day series (group E) and a 100-day series (group F) and of 6 rabbits of a 212-day series (group G) with experimentally induced unilateral hematogenous obstructive E . coli pyelonephritis were histologically examined and the glomerular and tubular lesions quantitatively evaluated using an ocular micrometer . An attempt was made to correlate the morphological changes with the results of simultaneous enzyme analyses (acid and alkaline phosphatases, glutaminase I).

Ann Neurol, 1977 Jul, 2(1), 49 - 56
Neonatal endotoxin encephalopathy; Gilles FH et al.; Telencephalic white matter of the neonatal kitten frequently contained diffuse astrogliosis or focal necrosis (sometimes including the thalamus and the caudate) following a single intraperitoneal injection of Escherichia coli lipopolysaccharide . No evidence for a disseminated intravascular coagulopathy was found . Telencephalic lesions in neonatal monkey and rabbit were also hemorrhagic . Enhanced karyorrhexis of glial nuclei was presented in the telencephalic white matter of the neonatal rat . In the kitten, a delay in the generation of macrophages and hypertrophic astrocytes occurs following transient neonatal endotoxemia . Marked weight loss and temperature fluctuation are prominent systemic effects . Large hemispheric cavitary lesions are not accompanied by obvious neurological deficits in the kitten.

J Biochem (Tokyo), 1977 Jul, 82(1), 261 - 6
Use of rabbit antiboty IgG bound onto plain and aminoalkylsilyl glass surface for the enzyme-linked sandwich immunoassay; Kato K et al.; Rabbit antibody IgG was bound onto aminoalkylsilyl or plain glass rods by simple adsorption . For comparison, rabbit antibody IgG was also bound onto glutaraldehyde-activated aminoalkylsilyl glass rods . These antibody-glass rods were tested by the sandwich procedure using Fab' fragments of rabbit antibody conjugated with beta-D-galactosidase from Escherichia coli . The glutaraldehyde-activated aminoalkylsilyl glass showed the largest capacity to bind antigen and the plain glass showed the smallest . However, the antibody-glass rods prepared by simple adsorption were as useful for the sandwich immunoassay of macromolecular antigens as those prepared with glutaraldehyde . With all the antibody-glass rods prepared, 0.1 to 10 fmol of ornithine delta-aminotransferase from rat liver and 2,4-dinitrophenyl human IgG were measurable . More than 10 fmol of the antigens may be measurable with larger amounts of the antibody-beta-D-galactosidase complexes, although the non-specific binding of the complexes to the solid phase increases to limit the sensitivity of the immunoassay.

Appl Environ Microbiol, 1977 Jul, 34(1), 18 - 22
Growth kinetics of Colpoda steinii on Escherichia coli; Drake JF et al.; Colpoda steinii was grown in two-stage continuous cultures with Escherichia coli as prey species . The concentration of prey and the ciliate mean cell volume, dry weight, and number per milliliter were determined at known growth rates . Steady states were reached in the second-stage continuous cultures at all growth rates . Although changes occurred in mean cell size of the ciliates and in the number per milliliter at various growth rates, the yield of protozoan biomass per unit of prey consumed was constant at all growth rates . The data were compared with several equations proposed to describe the kinetics of protozoan growth as a function of prey density.

Mol Biol (Mosk), 1977 Jul-Aug, 11(4), 917 - 32
{Circular dichroism of DNA--dye complexes . II . Anisotropy of the long-wave circular dichroism effect and structure of the complex}; Poletaev AI et al.; Anisotropy of torsional strength of the splitted electronic transition in the case of chromophore-chromophore interaction of dye molecules situated on the helical matrix was considered theoretically and as analytical expression for the value Rperpendicular/Rparallel was obtained . These theoretical results were compared with the experimental data obtained with DNA-proflavine, DNA-pyronine and DNA-acridine orange complexes oriented in multicappilar flow-cell . Studies of the optical effects (optical density and CD changes) due to orientation of these complexes showed that the acridine chromophores are not perpendicular with respect to the DNA axis (alpha D = 19--22 degrees) . The DNA base pairs in complexes as assumed also are not perpendicular to the DNA axis, the inclination angle of their transition moments (for the band near 260 nm) being bigger than that of dye chromophores (24 degrees) . These results indicate that under experimental conditions used by us no intercalation can be observed.

Mol Biol (Mosk), 1977 Jul-Aug, 11(4), 854 - 63
{ATP analogs in the RNA-polymerase reaction}; Aivazashvili VA et al.; The influence of some ATP analogs with the modified ribose residues on the transcription in vitro was studied . It was shown that all analogs are weak inhibitors competing with in RNA synthesis . Under certain conditions (ionic strength = 0.13, 25 degrees C) the only effective inhibitor of RNA synthesis 3'-O-methyl-ATP arrests reaction irreversibly . Perhaps as a result of its incorporation into the terminal position of the growing RNA chain . The increase of the temperature and of the ionic strength results in changes of the character of inhibition: 3'-O-methyl-ATP becomes (like other analogues) a reversible competitive inhibitor with a Ki = 4 . 10(-5) M . Some speculations concerning the mechanism of inhibition are discussed.

Gene, 1977 Jul, 1(5-6), 331 - 45
Isolation and characterization of the biotin genes of Escherichia coli K-12; Das Gupta CK et al.; DNA containing the biotin gene cluster, bioABFCD, of E . coli K-12 has been isolated from the EcoRI cleavage products of lambdabiot124-10 phage DNA and subsequently characterized by electron microscopic studies . The biotin-DNA fragment obtained after EcoRI cleavage of the lambdabiot124-10 DNA measures 18.7% lambda DNA length (approx . 9000 base pairs) . In addition to the biotin genes, it contains 4.75% and 3.08% lambda phage DNA at the left and right end-points of the bioABFCD cluster, respectively . The two bio promoter sites of the divergently transcribed biotin genes have been visualized under the electron microscope by binding RNA polymerase holoenzyme to the biotin DNA fragment . The two promoters are located at 41% and 43% length of the DNA fragment from its left endpoint . In vitro transcription of RNA from the bio-tin-DNA fragment has been visualized with the electron microscope, but so far no simultaneously transcribing "RNA:DNA" loops of the divergently oriented genes have been observed.

Gene, 1977 Jul, 1(5-6), 305 - 21
Cloning of chemically synthesized lactose operators; Sadler JR et al.; Recombinant DNA molecules, constructed from the ColE1-Mk5 hybrid plasmid PMB9 and a chemically synthesized wild-type lactose operator segment, have been used to transform Escherichia coli . Up to 10% of the transformants (selected for the tetracycline-resistance property of PMB9) are partially constitutive for the lactose operon enzyme beta-galactosidase . In vitro studies demonstrate that these partially constitutive transformants contain plasmid DNA molecules which carry one or more lactose operators, and which will bind purified lactose repressor . Preliminary results with some modified operator sequences are also presented.

J Microsc, 1977 Jul, 110(2), 121 - 32
Freeze-fracturing of monolayers (capillary layers) of cell, membranes and viruses: some technical considerations; Nermut MV et al.; A novel hinged device for freeze-fracturing of cell monolayer in the Balzers freeze-etch unit is described . It is economical on biological material and enables oriented adsorption of sheet-like membrane fragments . For freeze-fracturing 'by hand' a monolayer is formed on a positively charged piecie of mica (with polylysine) and this is covered with another piece of mica, thin brass plate of filter paper . Such a sandwich is frozen in liquid nitrogen and fractured by means of forceps . Several modifications of this technique as well as practical examples are described . Among possible application are: negative staining of intramembranous protein particles; chemical or physical analyses of single membrane leaflets; identification of protein complexes by immunoelectron microscopy, etc.

Zentralbl Bakteriol {Orig A}, 1977 Jul, 238(3), 350 - 4
{Heat-stable Escherichia coli-enterotoxin: reduced action after administration of phenylbutazone in infant mice (author's transl)}; Ohgke H et al.; Heat-stable Escherichia coli enterotoxin was assayed by the method of DEAN determining gut weight to body weight ratios in infant mice . The enterotoxic responses were significantly lower than in saline treated controls when Phenylbutazone at 20 mcg/mouse was administered subcutaneously 30 minutes prior to intragastric toxin challenge . The trials were performed with lyophilized culture filtrates of E . coli O 149:K91 (B) K88 ab (L), and three other enterotoxigenic strains one of which was isolated from an outbreak of swine oedema disease, the other two strains originated from the stools of diseased children.

Res Vet Sci, 1977 Jul, 23(1), 97 - 101
The effect of Freund's complete adjuvant on the cellular immune response in mice to a porcine strain of Escherichia coli lipopolysaccharide; Allan D et al.; The effect of Freund's complete adjuvant (FCA), known to enhance and prolong both cellular and humoral responses to thymus dependent (TD) antigens, was studied with regard to the cellular response in BALB/c mice to the thymus independent lipopoly-saccharide antigen of Escherichia coli O138, a porcine pathogen . Techniques based on immunocytoadherence (ICA), inhibition of ICA with an antiserum to the brain-associated theta alloantigen, immune adherence and macrophage migration inhibition, were used in this study . Apart from enhancing the rosette forming cell response, it is suggested that FCA appears to promote the action of the lipopolysaccharide on assembled macrophages with subsequent release of humoral factors which, in turn, activate T cells with consequent cell-mediated response.

Nucleic Acids Res, 1977 Jul, 4(7), 2511 - 26
Alteration of 5S RNA conformation by ribosomal proteins L18 and L25; Bear DG et al.; The effects of ribosomal proteins L18, L25 and L5 on the conformation of 5S RNA have been studied by circular dichroism and temperature dependent ultraviolet absorbance . The circular dichroism spectrum of native 5S RNA is characterized in the near ultraviolet by a large positive band at 267 nm and a small negative band at 298 nm . The greatest perturbation in the spectrum was produced by protein L18 which induced a 20% increase in the 267 nm band and no change in the 298 nm band . By contrast, protein L25 caused a small decrease in both bands . No effect was observed with protein L5 . Simultaneous binding of proteins L18 and L25 resulted in CD changes equivalent to the sum of their independent effects . The UV absorbance thermal denaturation profile of the 5S RNA L18 complex lacked the pre-melting behavior characteristic of 5S RNA . Protein L25 had no effect on the 5S RNA melting profile . We concluded that protein L18 increases the secondary, and possible the tertiary structure of 5S RNA, and exerts a minor stabilizing effect on its conformation while protein L25 causes a small decrease in 5S RNA secondary structure . The implications of these findings for ribosome assembly and function are discussed.

Nucleic Acids Res, 1977 Jul, 4(7), 2429 - 44
Hyperpolymer formation during renaturation of DNA from genomes with different sequence organisation; Flavell RB et al.; Hyperpolymer formation during the renaturation of DNAs from wheat, calf and E . coli was studied using hydroxyapatite chromatography, electron microscopy and S1 nuclease . Large hyperpolymers could not be eluted from hydroxyapatite with 0.5 M phosphate buffer at 60 degrees C . Large proportions of wheat and E . coli DNAs were incorporated into hyperpolymers when fragments 650 nucleotides long were renatured . A much smaller proportion of calf DNA was incorporated under equivalent conditions . Greater proportions of calf DNA accumulated in hyperpolymers only when longer fragments were incubated . Electron microscopy indicated no obvious differences in the basic structures of hyperpolymers formed by the three DNAs and confirmed the quantitative differences in hyperpolymer formation found by hydroxyapatite chromatography . It is concluded that the proportions and arrangement of the repeated sequences in the chromosomes of higher organisms determine the extent of rapid hyperpolymer formation during DNA renaturation in vitro.

Nucleic Acids Res, 1977 Jul, 4(7), 2389 - 406
Molecular cloning of extensive sequences of the in vitro synthesized chicken ovalbumin structural gene; Humphries P et al.; Double-stranded DNA molecules complementary to ovalbumin chicken messenger RNA were synthesized in vitro and integrated into the E . coli plasmid pCR1 using an oligodG-dc tailing procedure . The resultant hybrid plasmids, amplified by transfection of E . coli, were shown by hybridization and gel electrophoresis to contain extensive DNA sequences of the ovalbumin structural gene.

Nucleic Acids Res, 1977 Jul, 4(7), 2191 - 204
Chromatographic behavior of several mammalian tRNAs on acylated dihydroxyl-borate cellulose and Aminex A-28; Roe BA et al.; Studies of the chromatographic behavior of mammalian tRNAs, from several sources, on acylated DBAE-cellulose indicate that species of tRNA Asn , tRNA Asp and tRNA His can be retained on this matrix, while species of tRNA Tyr, tRNA Asn and tRNA Asp are not retained . Treatment of total rat liver tRNA with cyanogen bromide and subsequent chromatography on Aminex A-28 columns demonstrated that these tRNA species might contain Q (or Q*) nucleoside . However, comparable studies of the tRNA isolated from Walker 256 rat mammary tumor tissue demonstrated that this tumor tRNA almost totally lacks the hypermodified nucleosides Q and Q* . In addition, we have found that at least the major species of rat liver tRNA Asn contains the Q nucleoside . These studies indicate that chromatography on the acylated DBAE-cellulose matrix, couple with the analytical ion-exchange chromatography of cyanogen bromide treated and untreated amino-acyl-tRNA can be a valuable technique for the determination of alterations in the Q (or Q*) nucleoside content of the tRNAs isolated from normal and tumor tissues.

Nucleic Acids Res, 1977 Jul, 4(7), 2161 - 7
Covalent attachment of fluorescent probes to the X-base of Escherichia coli phenylalanine transfer ribonucleic acid; Schiller PW et al.; tRNA PheE, coli was labeled with the N-hydroxysuccinimide esters of 1-dimethylaminonaphthalene-5-sulfonyl glycine and N-methylanthranilic acid through reaction with the amino acid moiety of its X-base, whereby yields of 66% and 24%, respectively, were obtained . The purified dimethylaminonaphthalene-sulfonate derivative could not be aminoacylated and was found to be a strong competitive inhibitor of phenylalanine-tRNA synthetase {Ki=8X10(-7) M} . The N-methylanthraniloyl derivative could be charged to an extent of 5% as compared to native tRNA Phe . The fluorescence emission spectra of the derivatives are indicative of a slightly hydrophobic environment for both fluorophores . The results suggest that the integrity of the polar amino acid group of the X-base is required for the maintenance of the biologically active conformation.

Can J Comp Med, 1977 Jul, 41(3), 302 - 5
Intestinal emphysema (Pneumatosis cystoides intestinalis) in a gnotobiotic pig; Meyer RC et al.; A gnotobiotic pig monocontaminated with an enteropathogenic Escherichia coli was subsequently hyperimmunized to produce a monotypic antiserum . At necropsy, multiple, air filled cysts were found in the wall of the large intestine . The etiology of this condition is still conjectural . However, select strains of E . coli may cause or contribute to intestinal emphysema in swine.

Biokhimiia, 1977 Jul, 42(7), 1307 - 14
{In vivo and in vitro studies of phage T4 lysozyme mRNA translation}; Zaveniagina TN et al.; Translation of phage T4 lysozyme mRNA is studied in vivo and in vitro . Polyribosomes, carrying growing lysozyme polypeptides, are found to be homogenous enough and to contain 6 ribosomes . Complete molecules of phage lysozyme, which possess an enzymatic activity and are similar to the native enzyme in its electrophoretic mobility in polyacrylamide gel, have been synthetized in vitro on RNA isolated from phage-infected cells . The efficiency of RNA translation in cell-free system is discussed on the model of synthesis of functionally active individual protein.

Biofizika, 1977 Jul-Aug, 22(4), 640 - 5
{Interaction between a fluorescent probe and the surface structures of Escherichia coli}; Sabel'nikov AG et al.; The interaction of ANS with Escherichia coli cells, isolated cell walls, total cell envelopes and inner membranes was investigated in the presence and absence of Ca2+ ions . The addition of Ca2+ to intact cells at room temperature did not result in enhancement of fluorescence intensity . On the contrary the addition of Ca2+ to intact cells at 2 degrees C resulted in a progressive increase of fluorescence intensity with the maximum reached approximately by the 20th minute . The addition of Ca2+ to different membrane preparations showed a two-fold increase . Addition of Ca2+ to spheroplast membrane preparations did not result in any increase of the number of binding sites for ANS . There was a four-fold increase in quantum yields . All membrane preparations in titrations with Ca2+ showed saturation kinetics . The obtained results are discussed in terms of structural alterations in E . coli membranes and in relation with biological effects.

Proc Natl Acad Sci U S A, 1977 Jul, 74(7), 2830 - 4
Extracellular labeling of nascent polypeptides traversing the membrane of Escherichia coli; Smith WP et al.; To provide direct evidence for the hypothesis that secreted proteins may traverse membranes as growing chains, we labeled spheroplasts of Escherichia coli with a reagent (acetyl{35S}methionyl methylphosphate sulfone) that reacts with amino groups but does not cross the membrane . After fractionation, about 6% of the label in the membrane-polysome fraction was found to be attached to the polysomes . This attachment was via peptidyl-tRNA, as shown by several tests: release of most of the label from purified polysomes at low Mg2+; subsequent loss of about 25,000 daltons on cleavage by dilute alkali; release by puromycin; and release, accompanied by a marked increase in average molecular weight, on peptide chain completion . Moreover, a significant fraction of the completed chains was identified serologically and by molecular weight as a major periplasmic protein, alkaline phosphatase {orthophosphoric-monoester phosphohydrolase (alkaline optimum); EC 3.1.3.1} . This work provides direct evidence that: (i) secreted proteins thread through the membrane as growing peptide chains; and (ii) membrane-associated polysomes in bacteria are functionally attached to membrane and not merely trapped on disruption of the cell.

Proc Natl Acad Sci U S A, 1977 Jul, 74(7), 2756 - 60
Release of Escherichia coli DNA from membrane complexes by single-strand endonucleases; Abe M et al.; Treatment of gently prepared lysates of Escherichia coli with single-strand-specific endonuclease (SI or from mung beans) results in the release of about 90% of the DNA from membranes, as determined by the M band technique . The released DNA has an average molecular weight of about 1.2 X 10(8) . Data obtained with endonuclease S1 fit a mathematical model in which substrate sites are at or near membrane attachment sites . Data obtained with pancreatic deoxyribonuclease or x-rays fit a model for double-strand breaks at random sites along the DNA . Fitting data to these models, we estimate that there are 18+/-5 membrane attachment sites . The DNA remaining after S1 nuclease treatment is enriched for the region near the origin of chromosome replication . Therefore, attachment at this region near the origin of chromosome replication . Therefore, attachment at this region appears to be chemically different from that at the other sites along the DNA.

Proc Natl Acad Sci U S A, 1977 Jul, 74(7), 2710 - 4
Identification of spacer tRNA genes in individual ribosomal RNA transcription units of Escherichia coli; Morgan EA et al.; Transfer RNA genes ("spacer tRNA genes") are present in the spacer region between 16S and 23S rRNA genes in Escherichia coli . We have analyzed spacer tRNA genes carried by seven rRNA operons with different chromosomal locations . Six of these were isolated on plasmids and one on a transducing phage . We found that, in addition to the two previously identified genes for tRNA2Glu and tRNAIIle, there is a spacer tRNA gene which codes for tRNAIBAla . Of the seven rRNA operons studied, three had both tRNAIBAla and tRNAIIle genes, and the remaining four had the tRNA2Glu gene in their spacers . In addition, genes for tRNAIAsp were found near the distal ends of two different rRNA operons.

J Gen Microbiol, 1977 Jul, 101(1), 131 - 3
Pyrimidine dimer excision and DNA degradation during liquid holding recovery in ultraviolet-irradiated Escherichia coli K12 uvr+ rec; Masek F et al.; The survival of ultraviolet-irradiated Escherichia coli K12 uvr+re was increased by post-irradiation incubation in phosphate buffer . During this incubation both dimer excision and DNA breakdown were inhibited . It is suggested that the bacteria coped with the remaining dimers in a manner which did not involve excision.

J Gen Microbiol, 1977 Jul, 101(1), 112 - 30
Comparison of the flagellins from different flagellar morphotypes of Escherichia coli; Lawn AM; The molecular weights of the flagellins of 13 strains of Escherichia coli, each with a different H antigen, were estimated using polyacrylamide gel electrophoresis . In each case only one major polypeptide was demonstrated, although some strains possessed apparently sheathed flagella . Considerable differences in the molecular weight of flagellin accompanied the previously described structural differences between flagella from strains with different H antigens . The relationship between flagellar diameter and the molecular weight of the corresponding flagellins was similar for both unsheathed and apparently sheathed flagella . Crosss-polymerization occurred between seed consisting of fragment of unsheathed flagella and flagellin solution from apparently sheathed flagella and vice versa . Co-polymerization of flagellin from unsheathed flagella and flagellin from apparently sheathed flagella was also demonstrated . These polymerization experiments indicate that the assembly pattern of flagellin molecules is probably the same in all E . coli flagella . The above and other evidence suggests that there is no true sheath, but that the differences in flagellar surface structure between different E . coli flagella are the result of differences in the superficial parts of the flagellin molecules.

J Gen Microbiol, 1977 Jul, 101(1), 111 - 9
Morphological distinction between different H serotypes of Escherichia coli; Lawn AM et al.; The structure of the flagellar filaments of 50 Escherichia coli strains, each with a different H antigen, was examined . Although the flagella within each strain were structurally identical, there was variability in flagellar surface pattern between strains with differrent H antigens . Investigation of additional strains confirmed that flagella structure was the same in all strains having the same H antigen . In three pairs of strains with cross-reacting H antigens, the antigenic relatedness was associated with identical flagella structure.

Br J Pharmacol, 1977 Jul, 60(3), 471 - 6
Feline endotoxin shock: effects of methylprednisolone on kininogen-depletion, on the pulmonary circulation and on survival; Al-Kaisi N et al.; 1 Escherichia coli endotoxin, administered intravenously in a dose of 2 mg/kg to pentobarbitone anaesthetized, artificially ventilated cats resulted in pulmonary hypertension, systemic hypotension and an immediate (1-2 min) 30-40% reduction in plasma kininogen, an effect which probably indicates a release of plasma kinins . 2 Methylprednisolone (30 mg/kg), when administered 30 min before endotoxin, did not influence the endotoxin-induced pulmonary hypertension or systemic hypotension but completely prevented the depletion of plasma kininogen . 3 In spontaneously breathing cats, methylprednisolone, administered 30 min after endotoxin, caused a rapid repletion of kininogen and prolonged survival (47% at 6 h compared to 10% in the endotoxinalone animals) . Methylprednisolone did not appear to influence lactate production or the hyperventilation observed during the delayed endotoxin shock phase . 4 It is concluded t,at methylprednisolone does not prevent the release, by endotoxin, of a pulmonary vasoconstrictor prostaglandin, or its effects, but that perhaps by preventing kinin release it may reduce endotoxin-induced capillary leakage.

Br J Pharmacol, 1977 Jul, 60(3), 369 - 73
Anaphylactic reactions to endotoxin in guinea-pig tissues: relationship to endotoxin toxicity; McLean AJ; 1 A lipopolysaccharide extract of Escherichia coli 026:B6 cells (026:B6(B) endotoxin) was shown to be toxic to normal adult guinea-pigs . 2 The agent had no action on isolated preparations of ileum and heart taken from normal adult guinea-pigs . 3 Ileal segments from animals actively immunized against 026:B6(B) endotoxin showed dose-dependent contractions when exposed to endotoxin . Desensitization phenomena were demonstrated . 4 Reactivity of 026:B6(B) endotoxin was transferred to isolated preparations of ileum and heart from normal animals by passive transfer of immune serum . 5 Tissue responses to 026:B6(B) were associated with release of ileal spasmogen into the bath medium . Mepyramine blocked the effects of this spasmogen at bath concentrations which caused little change in ileal responses to carbachol . 6 It is concluded that E . coli endotoxin can elicit anaphylactic reactions, and that this process may potentiate endotoxin toxicity in sensitized animals . However, endotoxin toxicity in guinea-pigs does not appear to depend on this kind of allergic process.

Biull Eksp Biol Med, 1977 Jul, 84(7), 46 - 8
{Transformation of the substrate specificity of EcoRI restrictase under the influence of glycerin}; Karamov EV et al.; The restriction endonuclease EcoRI hydrolyzes DNA to a greater number of fragments in the presence of glycerol than under normal conditions . This enzyme begins to work by the so-called EcoRI-type of restriction when glycerol concentration reaches 50% . The EcoRI activity appeared in experiments only when the ionic strength of the solution was decreased and pH of the solution was increased . However, under such extreme conditions the enzyme was quickly inactivated and it was difficult to obtain reproducible results especially for hydrolysis of the high-molecular DNA . The suggested conditions for the EcoRI activity permit to obtain reproducible results, this being practically equivalent to discovery of the new restriction endonuclease.

Aviat Space Environ Med, 1977 Jul, 48(7), 654 - 8
Inefficiency of sanitation measures aboard commercial aircraft: environmental pollution and disease; Kikuchi R; Recent investigations at Tokyo International Airport have proven that environmental pollution resulting from the inefficient disposal of human excretion aboard aircraft is an important problem from the standpoint of quarantine . It is, therefore, recommended that the worldwide aviation industry take immediate measures to improve conditions and eliminate this problem, which has thus far been ignored by aircraft designers, airport administration, and CAB personnel.

Am J Trop Med Hyg, 1977 Jul, 26(4), 727 - 31
Immunosuppression mediated by adult worms in chronic schistosomiasis mansoni; Mota-Santos TA et al.; A marked reduction in the number of plaque-forming cells from spleens of mice infected with Schistosoma mansoni to sheep erythrocytes (SRBC) and lipopolysaccharide from Escherichia coli was observed . This reduction coincided with the late stages of the infection and was also observed in unisexual infection with male worms . Treatment of the animals with a schistosomicidal compound (oxamniquine) almost completely abolished the immunosuppression . The suppression could be induced by administration of 60 microgramg protein from worm membrane preparations (24 h before SRBC injection), but not by egg-extract injection . When the crude membrane preparation was injected 48 h before or 0 to 24 h after the SRBC challenge, the immunosuppression was not observed . Significant reduction of footpad swelling was also noted in infected mice when injected with SRBC.

Infect Immun, 1977 Jul, 17(1), 78 - 82
Diarrhea caused by Escherichia coli that produce only heat-stable enterotoxin; Levine MM et al.; To determine the role of Escherichia coli heat-stable enterotoxin (ST) as a virulence factor in human diarrhea, a strain that elaborates only ST (E . coli 214-4) was fed to free-living volunteers in doses of 10(6), 10(8), and 10(10) organisms . Short-lived (1 day) mild illness consisting of abdominal cramps with vomiting or diarrhea occurred in three of five individuals fed 10(8) . Typical travelers' diarrhea (loose stools, abdominal cramps, and low-grade fever for 2 to 3 days) was seen in four of five volunteers given 10(10); two had brief cholera-like purging of rice-water stools . Despite fever, there was no evidence of mucosal invasion . E . coli 214-4 became the predominant coliform in stools; coproculture isolates were uniformly negative for heat-labile enterotoxin (LT), whereas most produced ST . Ten of 13 individuals developed rises in antibody to somatic E . coli antigen, and none had rises in LT antitoxin . E . coli that elaborate only ST can cause diarrheal disease in adults.

Infect Immun, 1977 Jul, 17(1), 105 - 11
Patterns of loss of enterotoxigenicity by Escherichia coli isolated from adults with diarrhea: suggestive evidence for an interrelationship with serotype; Evans DJ Jr et al.; Enterotoxigenic Escherichia coli isolates obtained in Mexico from adult subjects with diarrhea and from healthy controls were examined for the production of heat-stable enterotoxin (ST) and heat-labile enterotoxin (LT) after serial passage in the laboratory . Isolates were found to be either stable for the production of ST and LT or unstable with respect to ST, LT, or both . Unilateral loss of either ST or LT production allowed classification of E . coli isolates into four groups according to stability/instability of enterotoxin production . Fewer serotypes, with more representative isolates, were in group I (stable) than in group IV (completely unstable) . Isolates from Dacca, Bangladesh, could be similarly classified into stability groups . There is an apparent relationship between serotype, stability of enterotoxin production, particularly LT, and isolation from diarrhea cases as opposed to isolation from healthy controls.

Cell, 1977 Jul, 11(3), 545 - 50
The araC promoter: transcription, mapping and interaction with the araBAD promoter; Hirsh J et al.; The start sites of the araC and araBAD gene messenger of E . coli were located by transcription in vitro from short DNA fragments, by high magnification electron microscopy and by genetic mapping . Transcription for these messengers proceeds in opposite directions from the start sites that are 150 base pairs apart . Transcription from the araBAD promoter requires araC protein plus arabinose and CAP protein plus cyclic AMP . In the experiments performed in vitro, inducing the araBAD promoter represses activity of the araC promoter.

J Immunol, 1977 Jul, 119(1), 65 - 72
Release of phospholipids from complement-mediated lesions on the surface structure of Escherichia coli; Inoue K et al.; When varying numbers of sensitized, 14C-labeled bacteria were treated with a certain amount of complement, in a fixed reaction volume, 14C compounds were liberated into the surrounding medium in proportion to the number of the bacteria, whereas the amount of the phospholipids liberated was constant regardless of the number of the bacteria even in the range of relative excess of complement . Since it is conceivable that a certain amount of complement might form a fixed number of lesions on the surface of all the sensitized bacteria, the amount of the liberated phospholipids seems to be proportional to the number of complement lesions . The 14C-materials released from complement-attacked bacteria were analyzed by isopycnic sucrose density gradient ultracentrifugation and they were mainly free phospholipids and other smaller molecules . A small amount of the smaller membrane proteins were also released as revealed by acid and SDS-polyacrylamide gel electrophoresis . These results suggest that the release of phospholipids is due to the displacement of membrane lipids by the complexes of the late acting complement components during their insertion into the membrane lipid bilayer.

J Immunol, 1977 Jul, 119(1), 263 - 70
Identification of RNA as a complement inhibitory component in an extract of Ehrlich ascites tumor cells; Renk CM et al.; A factor capable of inhibiting complement was obtained from intact Ehrlich ascites tumor cells by mild extraction with phosphate-buffered saline (PBS) . The inhibitor caused a decrease in extent of lysis of EAC14 with a concomitant extension of Tmax . EA, EAC1, EAC4 and EAC142 were all less susceptible to complement-mediated lysis after treatment with the tumor cell extract . Partial purification of a complement inhibitor was accomplished . The inhibitor was rich in RNA and its activity was totally destroyed by RNAase but not DNAase . RNA from mouse tissues, yeast, and Escherichia coli also inhibited complement hemolytic activity . The partially purified material only inhibited lysis of EAC1 and EAC14 . Slow inhibition of fluid phase C1 was also demonstrated . In addition, RNA-rich partially purified tumor cell extract was capable of precipitating with purified human C1q.

J Bacteriol, 1977 Jul, 131(1), 76 - 81
R-plasmid transfer and its response to nalidixic acid; Burman LG; The conjugational transfer efficiency of 41 wild-type R-plasmids was studied in Escherichia coli K-12 . Type I R-plasmids were transferred at comparatively high and rather uniform frequencies, whereas type F R-plasmids showed less uniform and, on average, somewhat lower transfer frequencies . R-plasmids not mediating sensitivity to F-, I-, or N-specific phages showed moderate transfer frequencies, and type N R-plasmids showed very low transfer frequencies . Various lines of evidence suggest that a well-expressed, but functionally inefficient, conjugation apparatus is the cause of the poor transfer of type N R-plasmids in liquid medium . Nalidixic acid efficiently inhibited transfer of type I and particularly type F R-plasmids, whereas the transfer of type N plasmids was resistant to the drug . Type F and type I plasmids appear to depend on at least one host function for their transfer, namely, the nalidixic acid-sensitive reaction in vegetative chromosome replication, whereas type N plasmids are independent of this function.

J Bacteriol, 1977 Jul, 131(1), 49 - 56
Fine-structure mapping and complementation analysis of the Escherichia coli cysB gene; Tully M et al.; Sixty-two point mutations were isolated in Escherichia coli by means of transduction with mutagenized phage P1 . Twenty-two deletions extending into cysB but able to recombine with at least some of the point mutations were isolated on a transmissible E . coli plasmid . Mapping of the point mutations against the deletions divided the former into 16 deletion groups . Nine merodiploids were constructed in which the chromosome carried one of the three point mutations most distal to the trp operon and in which a plasmid carried one of the three point mutations most proximal to the trp operon . All of these showed a Cys-phenotype . It follows that mutations at the two extreme ends of the region belong to the same complementation group.

J Bacteriol, 1977 Jul, 131(1), 372 - 3
Do cytochromes function as oxygen sensors in the regulation of nitrate reductase biosynthesis?
MacGregor CH, Bishop CW.
The observation that oxygen represses nitrate reductase biosynthesis in a hemA mutant grown aerobically with or without delta-aminolevulinic acid indicates that cytochromes are not responsible for nitrate reductase repression in aerobically grown cells.

J Bacteriol, 1977 Jul, 131(1), 347 - 55
Identification of lipopolysaccharides and phospholipids of Escherichia coli in polyacrylamide gels; Bailey SC et al.; Polyacrylamide gel electrophoresis of unfractionated lysates of radioactively labeled cells resolves not only proteins and polynucleotides into discrete bands but also cellular lipopolysaccharides and phospholipids . This allows a determination of the intracellular amounts of all of these macromolecules . In addition, this technique is sensitive enough to detect mutational alterations in lipopolysaccharide structure . Polyacrylamide gel electrophoresis is herein shown to be a useful tool for investigations into the structure of lipopolysaccharides and the synthesis of lipopolysaccharides and phospholipids.

J Bacteriol, 1977 Jul, 131(1), 30 - 3
Genetics of the relB locus in Escherichia coli; Diderichsen B et al.; A mutant of Escherichia coli with a delayed relaxed phenotype very similar to that of a previously described relB mutant has been obtained using a new selection procedure . The mutation giving rise to this phenotype has been shown to map at 34.5 min and to be 12% cotransducible with man . It is recessive, revertible, and most likely an allele of the relB gene.

J Bacteriol, 1977 Jul, 131(1), 270 - 9
Morphological analysis of the division cycle of two Escherichia coli substrains during slow growth; Woldringh CL et al.; Morphological parameters of the cell division cycle have been examined in Escherichia coli B/r A and K . Whereas the shape factor (length of newborn cell/width) of the two strains was the same at rapid growth (doubling time, tau, less than 60 min), with decreasing growth rate the dimensions of the two strains did change so that B/r A cells became more rounded and B/r K cells became more elongated . The process of visible cell constriction (T period) lasted longer in B/r A than in B/r K during slow growth, reaching at tau = 200 min values of 40 and 17 min, respectively . The time between termination of chromosome replication and cell division (D period) was found to be longer in B/r A than in B/r K . As a result, in either strain completion of chromosome replication seemed always to occur before initiation of cell constriction . Nucleoplasmic separation did not coincide with termination as during rapid growth but occurred in both strains within the T period, about 10 min before cell division.

J Bacteriol, 1977 Jul, 131(1), 214 - 23
Metabolism of arginine-specific messenger ribonucleic acid in Escherichia coli K-12; Natter W et al.; Ribonucleic acid-deoxyribonucleic acid (RNA-DNA) hybridization was employed for the determination of the level of messenger RNA (mRNA) transcribed from seven of the nine genes of the arginine regulon of Escherichia coli K-12 . The quantity of RNA complexing with each of the separated DNA strands of the argA, argF, argE, and argCBH operons carried on specialized transducing phages was measured . The derepressed:repressed ratio of mRNA formed in vivo was found to vary between about 3 and 4 when measured by hybridization to DNA isolated from specialized transducing phages carrying the argA, argE, argCBH, argF, and argI operons.

J Bacteriol, 1977 Jul, 131(1), 208 - 13
Effect of transient lambda prophage induction on ultraviolet light resistance and recombination in Escherichia coli; Braun A et al.; Transient induction of lambda prophage increases the ultraviolet light resistance of most exponentially growing Escherichia coli lysogens . Resistance is increased in wild-type, recB, recB recC, recB recC recF, and recB recC recL hosts . No enhancement in recA lysogens was found, nor was there enhancement in stationary cultures . Enhancement was dependent upon the lambdared recombination system . Transient induction also increases the genetic recombination rate in recB lysogens as measured in Hfr X F- matings.

J Bacteriol, 1977 Jul, 131(1), 153 - 62
Inactivation and partial degradation of phosphoribosylanthranilate isomerase-indoleglycerol phosphate synthetase in nongrowing cultures of Escherichia coli; Mosteller RD et al.; The stability of tryptophan biosynthetic enzyme activities was examined in cultures of repressor-negative (trpR) strains of Escherichia coli K-12 incubated under conditions of nutrient starvation of chloramphenicol inhibition . The results show that four of the five activities examined are stable under most nongrowing conditions, whereas one activity, indoleglycerol phosphate (InGP) synthetase, carried by the trpC protein, is unstable under most conditions tested . Phosphoribosylanthranilate (PRA) isomerase activity, which is also carried by the trpC protein, is unstable during starvation for ammonium, cysteine, or sulfate but is stable under other nongrowing conditions where InGP synthetase is not . InGP synthetase activity but not PRA isomerase activity is also diminished about twofold in cultures using glycerol as a carbon-energy source . These results indicate that one or both activities of the trpC protein is specifically inactivated under several culture conditions . Experiments with antibodies to the trpC protein show that sulfate-starved and ammonium-starved cultures contain 20 to 40% less immunologically reactive trpC protein than unstarved cultures . This indicates that the trpC protein is probably partially degraded under these conditions . During recovery from sulfate starvation or ammonium starvation, cultures slowly regain normal levels of InGP synthetase and PRA isomerase activities, suggesting that inactivation may be reversible.

J Bacteriol, 1977 Jul, 131(1), 145 - 52
Properties of the relaxation complex of supercoiled deoxyribonucleic acid and protein of R plasmid NR1 in Escherichia coli; Womble DD et al.; Some properties of the supercoiled deoxyribonucleic acid (DNA)-protein relaxation complex of the R plasmid NR1, which contains more than one origin for DNA replication, were examined . The percentage of complexed NR1 molecules that can be converted to the relaxed (nicked) form appeared to be unaffected by the conditions under which the host cells were cultured . However, the percentage of supercoiled NR1 DNA that can be relaxed was highly dependent on the method used to prepare the DNA and the agents used to induce relaxation . Our data suggest that 100% of NR1 molecules may exist in situ as DNA-protein relaxation complexes . An RTF-Tc segregant of NR1, which has deleted the r-determinants component of the NR1 and therefore does not contain the two origins of replication located in the r-determinants, has indistinguishable relaxation properties in comparison with NR1 itself.

J Bacteriol, 1977 Jul, 131(1), 105 - 10
Characterization of methylated neutral amino acids from Escherichia coli ribosomes; Chang FN et al.; The methylated neutral amino acids from both 30S and 50S ribosomal subunits of an Escherichia coli K strain were characterized . The 50S ribosomal subunit contains three methylated neutral amino acids: N-monomethylalanine, N-monomethylmethionine, and an as yet unidentified methylated amino acid found in protein L11 . Both N-monomethylalanine and N-monomethylmethionine were found in protein L33 . The amount of N-monomethylmethionine in this protein, however, is variable but not more than 0.25 molecules per protein . Thus protein L33 from this E . coli K strain has heterogeneity in its N-terminal amino acid and can start with either N-monomethylalanine or N-monomethylmethionine . The N-monomethylmethionine residue was not derived from the reduction of N-formylmethionine in the protein . The 30S ribosomal subunit contains only one methylated neutral amino acid: N-monomethylalanine.

Clin Exp Immunol, 1977 Jul, 29(1), 122 - 31
Suppressor cells and loss of B-cell potential in mice infected with Trypanosoma brucei; Corsini AC et al.; The functional changes in splenic lymphoid populations from mice infected with T . brucei strain S42 were studied throughout the 3 weeks of infection . Within a week of infection, proliferation of B and T cells profoundly increased as shown by 3H-labelled thymidine incorporation and fluorescent staining of surface Ig; the spleen cells secreted high levels of both IgM and IgG immediately cells were put into culture; but with progressing infection this Ig production declined . The early effect on T cells was reflected by lack of responsiveness to PHA . B-cell potential was studied in low-density cultures treated with lipopolysaccharide (E . coli) . Normal spleen cells proliferate extensively in these cultures with subsequent secretion of IgG as well as IgM . The ability to proliferate and produce Ig in response to LPS was severely depressed by day 7 and almost totally absent by day 12 of infection . Removal of T cells from the spleen cells obtained early in infection partly restored the response to LPS but as the infection neared its fatal end, B-cell potential appeared to become exhausted . Macrophages obtained from infected mice even early in infection profoundly depressed the ability of normal spleen cells to proliferate and secrete immunoglobulin in LPS cultures . The general immunodepressing effect of trypanosomes can be attributed to clonal exhaustion of B-cell potential caused by an undefined blastogenic stimulus from the parasites which may operate at least in part by the generation of suppressive T cells and macrophages.

Proc Natl Acad Sci U S A, 1977 Jul, 74(7), 2720 - 4
A DNA fragment containing the origin of replication of the Escherichia coli chromosome; Marsh RC et al.; A 38 kilobase pair region of the Escherichia coli K12 chromosome containing the replication origin has been physically mapped with restriction endonucleases EcoRI and HindIII . Replication starts within or very near a 1.3 kilobase pair HindIII fragment in the middle of this region and proceeds outward in both directions with apparently equal speed . This pattern was observed in both dnaA and dnaC temperature-sensitive (ts) initiation mutants at the start of the synchronous round of replication which occurs after downshift from the nonpermissive to the permissive temperature.

Nucleic Acids Res, 1977 Jul, 4(7), 2455 - 66
Influences of amino acid, ATP, pyrophosphate and tRNA on binding of aminoalkyl adenylates to isoleucyl-tRNA synthetase from Escherichia coli MRE 600; Flossdorf J et al.; Aminoalcohol-AMP esters, structurally related to the assumed intermediates of the amino acid activation reaction, behave as competitive inhibitors both with respect to the amino acid and ATP, when tested in the ATP-(32P) PPi-exchange or the tRNA-charging reaction . However, closer investigation of the binding of norvalinyl adenylate to isoleucyl-tRNA synthetase from Escherichia coli MRE 600 by an equilibrium method shows that only the amino acid is a true competitor, while ATP cannot displace the ester from binding . Pyrophosphate enhances the stability of the ester-enzyme complex whereas tRNA is without detectable influence.

Proc Natl Acad Sci U S A, 1977 Jul, 74(7), 2958 - 62
Specific action of T4 endonuclease V on damaged DNA in xeroderma pigmentosum cells in vivo; Tanaka K et al.; The specific action of T4 endonuclease V on damaged DNA in xeroderma pigmentosum cells was examined using an in vivo assay system with hemagglutinating virus of Japan (Sendai virus) inactivated by UV light . A clear dose response was observed between the level of UV-induce unscheduled DNA synthesis of xeroderma pigmentosum cells and the amount of T4 endonuclease V activity added . The T4 enzyme was unstable in human cells, and its half-life was 3 hr . Fractions derived from an extract of Escherichia coli infected with T4V1, a mutant defective in the endonuclease V gene, showed no ability to restore the UV-induced unscheduled DNA synthesis of xeroderma pigmentosum cells . However, fractions derived from an extract of T4D-infected E . coli with endonuclease V activity were effective . The T4 enzyme was effective in xeroderma pigmentosum cells on DNA damaged by UV light but not in cells damaged by 4-nitroquinoline 1-oxide . The results of these experiments show that the T4 enzyme has a specific action on human cell DNA in vivo . Treatment with the T4 enzyme increased the survival of group A xeroderma pigmentosum cells after UV irradiation.

J Pediatr, 1977 Jul, 91(1), 65 - 8
Enteropathogens associated with pediatric diarrhea in Mexico City; Evans DG et al.; Enteropathogens were investigated as possible agents in pediatric diarrhea occurring in Mexico City during the summer of 1975 . Pathogens were identified in 47 (76%) of 62 cases . Rotavirus particles were detected in 16 cases . Enterotoxigenic Escherichia coli was detected in 29 cases; 11 were positive for heat-labile enterotoxin and 18 were positive for only the heat-stable form of enterotoxin . Multiple pathogens were found simultaneously in 15 (24%) of the study population . This study indicates that the etiology of pediatric summertime diarrhea in Mexico City is diverse . ETEC and RV were the most frequently encountered pathogens, yet they frequently occurred together and with other pathogens . ST-only strains of toxigenic E . coli were as frequently recovered as LT-E . coli suggesting that both forms of ETEC must be sought in future field studies.

J Biochem (Tokyo), 1977 Jul, 82(1), 311 - 4
Studies on the turnovers in vivo of adenosine di- and triphosphates in a coupling factor of Escherichia coli; Maeda M et al.; The metabolic stabilities of bound adenine nucleotides in a membrane-bound ATPase (EF1) {EC 3.6.1.3} of Escherichia coli were studied by estimating their rates of turnover in vivo . Two-thirds of the bound ATP prelabelled with 32Pi in EF1 molecules was retained after 3 h in a chase medium . The bound ADP was chased rapidly with a half time of decrease of less than 1 h, the rate similar to that of cytoplasmic free nucleotides . These results suggest that bound ATP in the EF1 is not a direct intermediate in oxidative phosphorylation.

Surgery, 1977 Jul, 82(1), 68 - 73
Prevention of endotoxin-induced changes in oxidative phosphorylation in hepatic mitochondria; DePalma RG et al.; E . coli endotoxemia affects hepatic energy linked function by uncoupling oxidation from phosphorylation . This study was done to determine whether a steroid, methylprednisolone sodium succinate (MPS), as well as excess substrate sodium succinate (SS), alters directly the effects of endotoxin on hepatic mitochondria . An assay system using alpha-ketoglutarate (alpha-Kg) was developed to test this hypothesis . Isolated rat hepatic mitochondria were first incubated in concentrations of MPS, ranging from 2.0 to 6.0 mg/ml . At these concentrations uncoupling identical to that occurring with addition of endotoxin resulted . However, a more dilute solution of MPS, 0.12 mg/ml, permitted normal mitochondrial function . Preincubation of MT in 0.12 mg/ml of MPS, as well as with sodium succinate, prevented endotoxin-induced uncoupling . Both endotoxin and steroid resulted in increased ATPase activity in the medium . While preincubation with MPS blocks the endotoxin effect, very high steroid concentrations alone are harmful . A direct action of steroids on mitochondria is evident, as well as a weaker protective effect due to excess substrate (alpha-Kg + SS) . Since mitochondria are probably in direct communication with extracellular fluid, the assay system permits interaction of endotoxin, steroids, and substrates which mimic those which occur in vivo . The results of this study account for the previously reported variable effects obtained when steroids have been tested in vivo.

J Bacteriol, 1977 Jul, 131(1), 331 - 9
Coordinate regulation by iron of the synthesis of phenolate compounds and three outer membrane proteins in Escherichia coli; McIntosh MA et al.; The biosynthesis of the low-molecular-weight iron carrier enterochelin and of three outer membrane polypeptides appears to be coordinately regulated by the amount of cell-associated iron in Escherichia coli K-12 . Measurements of iron acquisition made throughout the growth cycle in iron-deficient media indicate that a very rapid accumulation of iron occurs in the first 2 h of growth; there is comparatively little iron uptake during exponential growth, which results in a gradual decrease in the cellular iron content with each generation . When this level falls below 400 ng of iron per mg (dry weight) of cells, there is a simultaneous onset of synthesis of the three outer membrane polypeptides and of enterochelin . This coordinate regulation was also observed in cells able to transport iron actively using only citrate as an iron-carrier.

J Bacteriol, 1977 Jul, 131(1), 229 - 39
Formation of sugar phosphates in colicin K-treated Escherichia coli; Takagaki Y et al.; Colicin K greatly decreased the incorporation of 32P-labeled inorganic orthophosphate into nucleotides and nucleic acids, causing a concomitant increase in the formation of 32P-labeled sugar phosphates in sensitive cells of Escherichia coli . These sugar phosphates were formed in aerobically growing cells, as well as in cells under stringent control of ribonucleic acid synthesis . The main 32P-labeled product was identified as sedoheptulose 7-phosphate in two strains (B1 and K-12 MK-1) and fructose 1,6-diphosphate in one strain (K-12 CP78) . The formation of sugar phosphates induced by colicin K was inhibited by carbonyl cyanide m-chlorophenylhydrazone . It was also not observed in N,N'-dicyclohexylcarbodiimide-treated cells or Mg2+-(Ca2+)-adenosine triphosphatase-less mutant (strain K-12 AN120) cells . Thus, the formation of sugar phosphates in colicin K-treated cells is dependent on the formation of adenosine 5'-triphosphate by oxidative phosphorylation.

Infect Immun, 1977 Jul, 17(1), 205 - 14
Mild alkaline hydrolysis of lipopolysaccharide endotoxin enhances its mitogencity for murine B cells; Goodman GW et al.; Mild alkaline hydrolysis was found to enhance the mitogenicity of lipopolysaccharide endotoxin for murine B lymphocytes . Alkaline treated lipopolysaccharide also retained its property as a polyclonal activator . Whereas this treatment reduced the lethality of endotoxin for mice, its toxicity for lymphocytes cultured in the absence of fetal calf serum was increased . Lipid analysis indicated that there were no significant changes in the fatty acids of lipid A, but particle size was significantly reduced and the material was more homogeneous and soluble than untreated lipopolysaccharide . The relationship of these effect on the structure of lipopolysaccharide endotoxin to the mechanism of B-lymphocyte activation is discussed.

C R Acad Sci Hebd Seances Acad Sci D, 1977 Jun 27, 284(24), 2569 - 72
{4-Thiouridine and photoprotection in Escherichia coli K 12}; Thomas G et al.; A high level of protection is observed in the Escherichia coli K 12 strain AB 1157 rec A 1 nuv+ whose transfer RNA contains 4-thiouridine . In contrast, the photoprotection level is low and observed at higher doses in a strain which differs from the former by a single mutation, nuv-, (lack of 4-thiouridine) . This nucleoside is therefore an important chromophore leading to photoprotection . This conclusion is corroborated by the similarity of the action spectra for 8-13 link formation in tRNA and for photoprotection.

J Biol Chem, 1977 Jun 25, 252(12), 4418 - 20
19F nuclear magnetic resonance of 5-fluorouridine-substituted tRNA1Val from Escherichia coli; Horowitz J et al.; The 19F NMR spectrum of Escherichia coli tRNA1Val in which {5-19F}uridine replaces 93% of all uridine and uridine-derived residues has been examined at 93.6 and 235 MHz . The resolution of 11 peaks and visibility of two additional shoulders at either frequency for the 14 FUra residues in the molecule attests to the excellence of 19F as a probe for the structure of tRNA1Val in solution . No significant gain in resolution was attained at the higher frequency . A comparison of the relative areas in the different regions of the 19F spectrum of mixed {FUra}tRNAs with that of {FUra}tRNA1Val suggests that the three single resonances at lowest field in the region 86.5 to 88.5 ppm upfield from trifluoroacetate correspond to the three invariant bases which form tertiary hydrogen bonds in all tRNAs, namely, 8 (U or s4U), 54 (T), and 55 (phi) in unsubstituted tRNAs.

J Biol Chem, 1977 Jun 25, 252(12), 4151 - 6
Charges of nicotinamide adenine nucleotides and adenylate energy charge as regulatory parameters of the metabolism in Escherichia coli; Andersen KB et al.; Methods for measurements of catabolic reduction charge (defined as NADH/(NADH+NAD+)) and anabolic reduction charge (defined as NADPH/(NADPH + NADP+)) are described using {14C}nicotinamide labeling of Escherichia coli cultures . Together with these parameters the adenylate energy charge (ATP + 1/2ADP)/(ATP + ADP + AMP) was measured using labeling with {2-3H}adenine . These three charges were found under different exponential growth conditions to have values independent of the growth conditions: catabolic reduction charge, 0.05; anabolic reduction charge, 0.45; and adenylate energy charge, 0.9 . The charges were examined during interruption of growth primarily affecting catabolism, respiration, or anabolism, leading to changes of the charges . The changes of charges are evaluated as a possible regulation of the metabolic rates utilizing or producing the nucleotides by their respective charges.

Mol Gen Genet, 1977 Jun 24, 153(3), 325 - 9
Further evidence that the ribosomal 30S proteins S3, S5, S9, S11, S12, and S18 possess specific 16S RNA binding sites; Hochkeppel HK et al.; E . Coli ribosomal 16S RNA prepared by an acetic acid-urea extraction technique individually binds, in addition to the seven established proteins, 6 new 30S ribosomal proteins (S3, S5, S9, S12, S18 and S11) (Hochkeppel et al., 1976) . In this communication we demonstrate the site specificity of these proteins . Binding curves of the individual proteins with acetic acid-urea 16S RNA show that the binding of all six proteins to the RNA reaches a plateau at 0.3-0.97 copies per 16S RNA molecule . No significant binding of these proteins to classicial phenol extracted 16S RNA is observed, with the exception of S13 which binds 0.2 copies of protein per molecule of 16S RNA . Specificity of binding of these proteins is also demonstrated in "chase" experiments . The site specificity of individual {3H}-labeled 30S proteins bounds to 16S RNA is tested by the addition of non-radioactive 30S total protein to the reaction mixture.

Mol Gen Genet, 1977 Jun 24, 153(3), 289 - 95
Involvement of IS1 in the dissociation of the r-determinant and RTF components of the plasmid R100.1; Chandler M et al.; The formation of the r-determinant pLC1 and of the RTF pAR132 from the composite plasmid R100.1 was investigated . The general location of IS1 sequences on the three plasmids was established by hybridization of lambdar14 CII::IS1 DNA to EcoRI generated fragments of the various plasmids separated by agarose gel electrophoresis and transferred directly to nitrocellulose filters . The position of IS1 sequences on these fragments and the homologies between fragments were analyzed by electron microscopy of heteroduplex molecules . The results show that the excision of both pLC1 and pAR132 occurred by an exchange between the two IS1 sequences present on R100.1.

Arch Microbiol, 1977 Jun 20, 113(3), 185 - 9
A continuous culture study of an ATPase-negative mutant of Escherichia coli; Stouthamer AH et al.; For anaerobic glucose-limited chemostat cultures of Escherichia coli a value of 8.5 was found for YmaxATP . For anaerobic glucose- or ammoniumlimited chemostat cultures of the ATPase-negative mutant M2-6 of E . coli YmaxATP values of 17.6 and 20.0 were found, respectively . From these data it can be concluded that in the wild type during anaerobic growth 51-58% of the total ATP production is used for energetization of the membrane . Using the YATP values obtained in the anaerobic experiments a P/O ratio of 1.46 could be calculated for aerobic experiments with the wild type . It is concluded that from the energy obtained by respiration in wild type E . coli about 60% is used for membrane energetization and only about 40% for the actual formation of ATP . No dramatic difference in the maintenance requirement for ATP or glucose has been observed between glucose- and ammonium-limited chemostat cultures of the mutant . The large difference in maintenance requirement observed for such cultures of the wild type is therefore supposed to be made possible by ATP hydrolysis by the ATPase.

Science, 1977 Jun 17, 196(4296), 1313 - 9
Rat insulin genes: construction of plasmids containing the coding sequences; Ullrich A et al.; Recombinant bacterial plasmids have been constructed that contain complementary DNA prepared from rat islets of Langerhans messenger RNA . Three plasmids contain cloned sequences representing the complete coding region of rat proinsulin I, part of the preproinsulin I prepeptide, and the untranslated 3' terminal region of the mRNA . A fourth plasmid contains sequences derived from the A chain region of rat preproinsulin II.

Biochim Biophys Acta, 1977 Jun 17, 476(4), 321 - 32
Genetic regulation of the constitutive D-ribose operon in Escherichia coli B/r; Abou-Sabe M et al.; Merodiploid complementation analysis of the constitutive synthesis of the D-ribokinase and the D-ribose permease in Escherichia coli B/r has shown that the constitutive D-ribose operon is genetically controlled by a transdominant regulatory gene closely linked to the D-ribokinase and D-ribose permease structural genes . The regulatory mechanism for this operon shows no requirement for operator-repressor interaction, rather a truly positive control mechanism and thus suggests an extension of the operon model in its application to constitutive enzyme regulation in bacteria.

Biochim Biophys Acta, 1977 Jun 16, 467(3), 386 - 95
Functional symmetry of the beta-galactoside carrier in Escherichia coli; Teather RM et al.; Cytoplasmic membrane vesicles with either normal or inverted orientation were prepared from Escherichia coli . The lactose transport activity of these vesicle preparations was compared . The parameters measured were net efflux, counterflux, and K+/valinomycin-induced active uptake of lactose . With membrane vesicles derived from both wild-type and cytochrome-deficient strains the right-side-out and inverted membrane preparations showed similar rates of lactose flux in all assays . According to these criteria, the activity of the beta-galactoside transport protein is inherently symmetrical . One major difference was observed between the native and inverted vesicle preparations: the inverted vesicles had approximately twice the specific activity of native vesicles in the counterflux and K+/valinomycin-induced uptake assays . This difference can be largely ascribed to the presence in the normal vesicle preparation of vesicles with a high passive permeability to lactose . Such vesicles are apparently absent from the inverted vesicle preparations.

Eur J Biochem, 1977 Jun 15, 76(2), 425 - 32
The recBC enzyme of Escherichia coli K12: premature cessation of catalytic activities in vitro and reactivation by potassium ions; Hermanns U et al.; It is shown that in vitro the degradation of native and single-stranded DNA as well as the hydrolysis of ATP by purified recBC enzyme ceases 2-3 min after the start of the reaction . The presence of potassium ions (60-100 mM), bovine serum albumin (1 mg/ml) or protein from cell-free Escherichia coli extract (10 microgram/ml) prevents the cessation of the activity . Once the cessation has occurred, the activity of the enzyme can be completely restored by the addition of potassium ions, but not by bovine serum albumin . Sedimentation studies revealed that, in contrast to the active recBC enzyme, the 'silent' enzyme is no longer associated with substrate DNA of high molecular weight . On the basis of these results and other observations it is hypothesized that during the degradation of DNA in the absence of potassium ions or bovine serum albumin the recBC enzyme is subject to an alteration of its molecular conformation which results in an inactive form.

Biochemistry, 1977 Jun 14, 16(12), 2800 - 5
Thermodynamic studies of the reversible association of Escherichia coli ribosomal subunits; Hui Bon Hoa G et al.; The kinetics of association of Escherichia coli 30S and 50S ribosomal subunits have been carried out as a function of temperature after a magnesium jump from 1.5 to 3 mM . Turbidimetric recordings combined with a stopped-flow apparatus were used to follow the kinetics . The data show that the rates of formation and dissociation of the 70S particles at 3 mM Mg2+ and +25 degrees C were, respectively: k2 = 10(5) M-1 s-1, k1 = 4,5 X 10(-3) s-1; lowering the temperature decreases the rate constants with activation energies equal to E2 = 7.5 kcal/mol, E1 = 26.5 kcal/mol and enhances the association equilibrium towards the 70S species with an enthalpy change (delta H degrees assoc = -19.9 kcal/mol) dominant over the entropy change (delta S degrees assoc = -33 cal/(deg mol)) . These thermodynamic parameters were compared to those obtained from studies on the interactions of codon-anticodon in yeast phenylalanine transfer RNA as well as of ribooligonucleotides . The kinetic and thermodynamic data are shown to be consistent with 16S-23S RNA interaction.

Mol Gen Genet, 1977 Jun 8, 153(2), 185 - 90
Repression of inducible enzyme synthesis in a mutant of Escherichia coli K 12 deleted for the ptsH gene; Gershanovitch VN et al.; The genome of lambda phage with thermosensitive repressor was inserted into the pts region of the Escherichia coli chromosome . This lysogenic culture possessed the PTS1 phenotype at 30 degrees C . A mutant strain with a deletion covering the ptsH gene was isolated after a prophage curing procedure . The deletion nature of the pts mutation was confirmed in genetical and biochemical experiments . The deletion covered a small fragment of the bacterial genome not extending in the ptsI and lig genes . The isolated deltaptsH mutant possessed all characteristics of known pts mutants: pleiotropical disturbances of transport and utilization of a number of carbohydrates, repression of the enzyme inducible synthesis, and resistance to glucose catabolite repression . From these and other data we can conclude that the phosphorylated form of the heat-stable protein HPr is involved (directly or indirectly) in activation of the DNA transcription process.

Mol Gen Genet, 1977 Jun 8, 153(2), 115 - 20
A new ribosomal protein locus in Escherichia coli: the gene for protein S6 maps at 97 min; Isono K et al.; A mutant of E . coli selected for temperature-sensitive growth on rich medium harbored an altered ribosomal protein S6 (Isono et al., 1976) . This mutant was found to possess at least two mutations, one being responsible for the temperature-sensitivity and the other for the S6 alteration . Crosses with various Hfr strains as well as transductions with P 1kc revealed that the former mutation mapped at 98 min and the latter at 97 min . Furthermore, rec A derivatives of this mutant heteromerodiploid for this region possessed both the wild type and themutant forms of S6 . Thus it was established that the gene at 97 min was indeed the structural gene for protein S6 (rpsF) and not a gene modifying it.

C R Acad Sci Hebd Seances Acad Sci D, 1977 Jun 6, 284(21), 2171 - 4
{Study of marine microvibrios of Roscoff (Microvibrio marinus roscoffensis)}; Guelin A et al.; Marine microvibrios, predators of E . coli, have been detected worldwide, from equatorial waters to the polar regions . The morphology and behavior of two microvibrios have been studied . Their multiplication and aggressiveness in regard to their bacterial prevy are not impeded by dialysis membranes and manifest themselves, away from all direct contact between the predator and its prey.

C R Acad Sci Hebd Seances Acad Sci D, 1977 Jun 6, 284(21), 2167 - 70
{Synthesis of specific proteins by the nucleoid of Escherichia coli}; Nisman B et al.; The induction of beta-galactosidase and alkaline phosphatase by the nucleoid of Escherichia coli was studied . Only the membrane-associated form was active in the presence of S 30 . The induction of beta-galactosidase showed an absolute requirement for the inducer and was enhanced by cyclic AMP and cyclic GMP . Further-more, in our hands, the synthetic activity of the membrane-associated nucleoid proved to be far higher than that of the soluble system described by Zubay . Our results suggest that membrane shield the structure which is necessary for the integrity of the initiation step of both transcription and translation.

Eur J Biochem, 1977 Jun 1, 76(1), 189 - 96
Characterisation of RNA fragments obtained by mild nuclease digestion of 30-S ribosomal subunits from Escherichia coli; Rinke J et al.; When Escherichia coli 30-S ribosomal subunits are hydrolysed under mild conditions, two ribonucleoprotein fragments of unequal size are produced . Knowledge of the RNA sequences contained in these hydrolysis products was required for the experiments described in the preceding paper, and the RNA sub-fragments have therefore been examined by oligonucleotide analysis . Two well-defined small fragments of free RNA, produced concomitantly with the ribonucleoprotein fragments, were also analysed . The larger ribonucleoprotein fragment, containing predominantly proteins S4, S5, S8, S15, S16 (17) and S20, contains a complex mixture of RNA sub-fragments varying from about 100 to 800 nucleotides in length . All these fragments arose from the 5'-terminal 900 nucleotides of 16-S RNA, corresponding to the well-known 12-S fragment . No long-range interactions could be detected within this RNA region in these experiments . The RNA from the smaller ribonucleoprotein fragment (containing proteins S7, S9 S10, S14 and S19) has been described in detail previously, and consists of about 450 nucleotides near the 3' end of the 16-S RNA, but lacking the 3'-terminal 150 nucleotides . The two small free RNA fragments (above) partly account for these missing 150 nucleotides; both fragments arose from section A of the 16-S RNA, but section J (the 3'-terminal 50 nucleotides) was not found . This result suggests that the 3' region of 16-S RNA is not involved in stable interactions with protein.

J Bacteriol, 1977 Jun, 130(3), 1234 - 43
Properties of an Escherichia coli K-12 mutant defective in the transport of arginine and ornithine; Celis TF; A canavanine-resistant mutant strain, defective in the transport of arginine and ornithine, was isolated and characterized . Experiments presented show that both the kinetics of influx and the steady state of accumulation of arginine and ornithine are affected by the mutation, whereas the activity of other related transport systems remains unchanged . On the basis of competitive studies, it is concluded that L-canavanine can inhibit efficiently the arginine-specific uptake system . D-Arginine appears to be a moderate inhibitor . None of the basic amino acid-binding proteins of the mutant strain showed detectable alterations in terms of quantity, physical properties, or affinity constants . Studies on the relationship between the number of transport carriers and the steady state of accumulation of arginine suggested the presence of a reduced number of membrane carriers in the mutant strain . It is proposed that the mutation affects a regulatory gene concerned with controlling the amount of membrane carriers produced, which are components of the arginine- and ornithine-specific uptake systems . The mutation maps at min 62 on the recalibrated linkage map of Escherichia coli K-12, in a locus closely linked or identical to argP.

Biken J, 1977 Jun, 20(2), 47 - 55
Factors affecting the formation of alkaline phosphatase isozymes in Escherichia coli K-12; Nakata A et al.; Physiological and genetic factors affecting the formation of isozymes of alkaline phosphatase in Escherichia coli K-12 were studied . Our results are compatible with the hypothesis proposed by Schlesinger and his co-workers (Schlesinger et al., 1975) that the multiple forms of the enzyme are produced by converting a newly synthesized one (the least negatively charged one) into less negatively charged forms . Neither energy source nor de novo synthesis of the enzyme was necessary for the conversion . It is also confirmed that the conversion is effectively inhibited by externally added arginine (Piggott et al., 1972) but only partially by canavanine (arginine analog) or casamino acids . We isolated a mutant strain which did not form isozyme(s), if any, under the condition in which the wild type strain formed isozymes . The mutation(s) was proved to be mapped in the locus (or loci) other than alkaline phosphatase structure gene in the E . coli genetic map . We tentatively proposed to designate this the iap gene(s), an abbreviation for isozyme of alkaline phosphatase, which plays a role in isozyme formation.

Southeast Asian J Trop Med Public Health, 1977 Jun, 8(2), 165 - 72
Intestinal and blood parasites in the North Lore District, Central Sulawesi, Indonesia; Carney WP et al.; Over 1,000 stool specimens from residents of the Napu and Besoa Valleys, Central Sulawesi, Indonesia were examined . Schistosoma japonicum was detected in 31% of Napu Valley residents while in only 2% of the Besoa Valley residents . Hookworm infections were the most frequently encountered helminth parasitisms in both valleys . Other helminth parasites encountered were: Ascaris lumbricoides, Trichuris trichiura, Enterobius vermicularis, Strongyloides stercoralis, Physaloptera sp., Diphyllobothrium sp., echinostome and heterophyid trematodes . Intestinal protozoa endemic to the area were: Entamoeba histolytica, E . coli, E . hartmanni, Iodamoebe butschlii, Giardia lamblia, Chilomastix mesnili and Trichomonas hominis . Plasmodium falciparum was responsible for malaria parasitaemias in 5% of 1353 specimens examined and Brugia malayi microfilaraemias were detected in 10% of 972 specimens examined.

Q Rev Biol, 1977 Jun, 52(2), 155 - 78
The biological effects and mode of action of L-canavanine, a structural analogue of L-arginine; Rosenthal GA; Many of the 200 or so non-protein amino acids synthesized by higher plants are related structurally to the constituents of common proteins . L-Canavanine, the guanidinooxy structural analogue of L-arginine, is representative of this group . It has provided valuable insight into the biological effects and the mode of action of non-protein amino acids which acts as analogues of the protein amino acids . The arginyl-tRNA synthetases of numerous canavanine-free species charge canavanine, and canavanine is subsequently incorporated into the nascent polypeptide chain . Production of canavanine-containing proteins ultimately can disrupt critical reactions of RNA and DNA metabolism as well as protein synthesis . Canavanine also affects regulatory and catalytic reactions of arginine metabolism, arginine uptake, formation of structural components, and other cellular precesses . In these ways, canavanine alters essential biochemical reactions and becomes a potent antimetabolite of arginine in a wide spectrum of species . These deleterious properties of canavanine render it a highly toxic secondary plant constituent that probably functions as an allelochemic agent that deters the feeding activity of phytophagous insects and other herbivores.

Nucleic Acids Res, 1977 Jun, 4(6), 2009 - 200
Rabbit liver tRNA1Val:II . unusual secondary structure of T psi C stem and loop due to a U54:A60 base pair; Jank P et al.; In contrast to all other known tRNAs, mammalian tRNA1Val contains two adenosines A59 and A60, opposite to U54 and psi 55 in the U psi CG sequence of the T psi C loop, which could form unusual A:U (or A: psi pairs in addition to the five "normal" G:C pairs . In order to measure the number of G:C and A:U (A: psi) pairs in the T psi C stem, we prepared the 30 nucleotide long 3'-terminal fragment of this tRNA by "m7G-cleavage" . From differentiated melting curves and temperature jump experiments it was concluded that the T psi C stem in this fragment is in fact extended by an additional A60:U54 pair . A dimer of this fragment with 14 base pairs was characterized by gel electrophoresis and by the same physical methods . An additional A:U pair in the tRNA1Val fragment does not necessarily mean that this is also true for intact tRNA . However, we showed that U54 is far less available for enzymatic methylation in mammalian tRNA1Val compared to tRNA from T-E . coli . This clear difference in U54 reactivity, together with the identification of an extra A60:U54 pair in the U psi CG containing fragment suggests the presence of a 6 base pair T psi C stem and a 5 nucleotide T psi C loop in this tRNA.

Nucleic Acids Res, 1977 Jun, 4(6), 1957 - 78
Use of specific endonuclease cleavage in RNA sequencing; Gupta RC et al.; Nonradioactive RNA fragments may be sequenced by incorporation of (3H)-label into 3'-terminal positions, controlled digestion with specific ribonucleases, and separation according to size of the digestion products on polyethyleneimine- (PEI-) cellulose thin layers . This combination of techniques allows one to measure accurately distances of specific cleavage sites from the labeled terminal positions . The cleavage specificities of RNases T1, U2, and A are utilized to identify the positions of G, A, and pyrimidine residues respectively . C and U may be distinguished by mobility differences on PEI-cellulose thin layers at ph 2.6 . The procedure is simple, rapid, and highly sensitive; as little as 0.5 - 1 microgram of a RNA of the size of tRNA will be needed to sequence all fragments in a complete RNase digest.

Nucleic Acids Res, 1977 Jun, 4(6), 1853 - 71
Alterations of tRNA modification in mammalian systems: the effect of ethionine; Friedman S; The relationship between the modification of tRNA and its ability to act as a substrate for homologous tRNA modification enzymes in vitro was studied . The tRNA extracted from the livers of rats was active as a substrate for in vitro methylation with extracts from normal rat liver 19 h after treatment with L-ethionine (35 mg/100 g/24 h) . After 4 weeks of feeding a diet containing o.25% DL-ethionine, the tRNA was a poor substrate for methylation in vitro, even though it was deficient in methylated nucleosides . Only 18% and 7% of the available sites could be methylated after 67 h and 4 weeks, respectively, of ethionine treatment . 3-(3-amino-3-carboxypropyl)uridine, a nucleoside that is also synthesized from S-adenosylmethionine, was assayed in individual tRNAs by their reactivity with the N-hydroxysuccinimide ester of phenoxyacetic acid . The reactivity of tRNAIle, tRNAAsn, and tRNAThr was decreased by treatment with ethionine at 67 h as well as at 2 and 4 weeks, although no difference could be detected at 19 h.

Nucleic Acids Res, 1977 Jun, 4(6), 1713 - 26
Inhibition of transcription of supercoiled PM2 DNA by carbodiimide modification; Flashner MS et al.; PM2 superhelican DNA (form I), which as been reacted with the single strand specific reagent, N-cyclohexyl-N'-beta-(methylmorpholinium)ethyl carbodiimide (CMC) is more than 95% inhibited in its ability to support transcription with E . coli B RNA polymerase in vitro . Almost complete inhibition of transcription was achieved after 2 hours of reaction with FI when only 1% of the bases were modified . A large increase in S20,* (from 26.8 S to 33.6 S) of FI DNA was observed during the course of reaction . Rifampicin resistant transcription is more susceptible to inhibition by CMC than total transcription, suggesting that the CMC is preferentially binding at promoter sites . These results clearly are in accord with the observation that supercoiled DNA contains localized regions of unpaired bases . The promotor sites for E . coli RNA polymerase in FI PM2 DNA appear to be located at or near these unpaired sites.

J Gen Microbiol, 1977 Jun, 100(2), 355 - 61
Genetic determinants of the synthesis of the polysaccharide capsular antigen K27(A) of Escherichia coli; Schmidt G et al.; Most of the his+ hybrids from crosses between the Escherichia coli donor Hfr45(O8:K27) and different E . coli O9 recipients expressed the donor O8 antigen specificity and produced the capsular antigen K27 . Therefore these hybrids must have inherited the his-linked donor rfb region determining the synthesis of O8- specific polysaccharides as well as his-linked genes involved in K27 antigen synthesis . In the living state these hybrids were inagglutinable in O8 antiserum like the donor cells . However, when E . coli K12 and O8:K42- were used as recipients most of the his+ hybrids were agglutinable in O8 and K27 antisera . The amounts of K27 antigen present in these hybrids, designated as K27i (intermediate) forms, were sufficient to evoke the production of K27 antibodies in rabbits, but insufficient to inhibit O-agglutination of the respective cells . The additional transfer of the trp region of E . coli O8:K27 into such K27i forms frequently resulted in O-inagglutinable K27+ hybrids . This is attributed to the introduction of trp-linked genes which apparently play a role in the synthesis of K27 capsular antigen . Tus it is concluded that at least two gene loci, one close to his and the other close to trp, are required for the synthesis of the complete capsular antigen K27.

Infect Immun, 1977 Jun, 16(3), 781 - 8
Humoral immune response to the heat-labile enterotoxin of Escherichia coli in naturally acquired diarrhea and antitoxin determination by passive immune hemolysis; Evans DJ Jr et al.; Acute- and convalescent-phase sera from 132 students attending a university in rural Mexico were assayed for antibody against the heat-labile enterotoxin (LT) of Escherichia coli by neutralization of LT activity in the Y-1 adrenal cell assay and by passive immune hemolysis of LT-sensitized sheep erythrocytes . The two titration methods produced comparable results with respect to antitoxin responses detected . An inverse relationship was found between acute geometric mean antitoxin titer and the occurrence of diarrhea associated with LT-producing E . coli, especially in newly arrived students from the U.S.A . A significant correlation (P less than 0.00 5) was found between a rise in antitoxin titer detectable by the passive immune hemolysis technique and diarrhea with LT-producing E . coli isolated . Thus, humoral antitoxin titers appear to be a useful indicator of immune status with respect to enterotoxigenic (LT) E . coli diarrhea.

Biophys J, 1977 Jun, 18(3), 245 - 67
A kinetic model of cooperativity in aspartate transcarbamylase; Dembo M et al.; A relatively simple kinetic model is proposed to account simultaneously for data on the binding of carbamyl phosphate and succinate to aspartate trans carbamylase (ATCase), and for the relaxation spectrum associated with this binding . The model also accounts for measurements of the initial velocity of the reaction of ATCase with respect to aspartate and carbamyl phosphate . The principal assumption made is that ATCase consists of three identical noninteracting cooperative dimers . Ordered binding and both sequential and concerted conformational changes in the dimers are needed to account for the properties of ATCase . The values of the parameters of this model can be determined by fitting to existing experimental evidence . Various new quantitative predictions are made that can serve as additional tests of the proposed theory.

Urology, 1977 Jun, 9(6), 667 - 9
Intrarenal mycotic aneurysm; Ganem EJ et al.; A case is presented of a sixty-seven-year-old man in whom Escherichia coli septicemia developed after catheterization, followed by left renal hemorrhage due to an intrarenal mycotic aneurysm . Because of clinical circumstances that prevailed, treatment was nonsurgical consisting of multiple blood transfusions and intravenous antibiotics . The aneurysm healed spontaneously as proved by subsequent renal arteriography.

Proc Natl Acad Sci U S A, 1977 Jun, 74(6), 2379 - 83
Nucleic acid helix-unwinding properties of ribosomal protein S1 and the role of S1 in mRNA binding to ribosomes; Kolb A et al.; The presence of ribosomal protein S1 in 30S ribosomes is indispensable for the formation of 30S initiation complexes with natural mRNA . The 30S subunits lacking S1 retain activity with AUG as mRNA and are also active in poly(rU)-directed binding of Phe-tRNA . Isolated protein S1 stoichiometrically disrupts the secondary structure of helical and stacked single-stranded polynucleotides and converts them into their fully or partially denatured forms . A mono-N-ethylmaleimide derivatives of S1 is nearly devoid of any RNA helix-unwinding properties but is readily incorporated into 30S subunits deficient in S1 . The resulting N-ethylmaleimide-S1-containing 30S subunits are completely inactive in the binding of MS2 {3H}RNA and in the formation of an initiation complex with MS2 RNA as mRNA . They retain activity in the binding of the initiator fMet-tRNA in response to the trinucleotide AUG and in the binding of Phe-tRNA in response to poly(U) . They also retain the capacity to bind 50S subunits and to form 70S couples . These results suggest that a correlation exists between the RNA helix-unwinding capacity of isolated S1 and the function of S1 in the ribosomal binding of natural mRNA when the protein becomes part of the 30S subunit.

Proc Natl Acad Sci U S A, 1977 Jun, 74(6), 2316 - 20
Shape of the 50S subunit of Escherichia coli ribosomes; Stuhrmann HB et al.; Extrapolation of a series of low-angle neutron scattering curves to infinitely high contrast gives a scattering function IC(kappa) which is dependent on the shape of the solute molecule . For the 50S subunit of E . coli ribosomes, the first part of the structure determination by neutron scattering, namely the determination of the molecular shape from IC(kappa), is reported . The result is in good agreement with models of the 50S subunit determined by electron microscopy.

Proc Natl Acad Sci U S A, 1977 Jun, 74(6), 2246 - 50
Experimental evidence for kinetic proofreading in the aminoacylation of tRNA by synthetase; Yamane T et al.; The enzymatic aminoacylation of tRNA can be viewed as a means of proofreading either the amino acid or the tRNA or both . We have conducted further experimental tests of kinetic proofreading in discriminating between cognate and noncognate amino acids and tRNAs as follows: (formula: see text) . In cases (i) and (ii) the amino acids are proofread, in cases (iii) and (iv) the tRNA is proofread, and in case (v), both the amino acid and the tRNA are proofread . ATP consumed per acylation was 400, 1.5, 40, 25, and 1000, respectively . High ATP/aminoacylation ratios are diagnostic for kinetic proofreading.

Eur J Biochem, 1977 Jun 1, 76(1), 51 - 61
A trypsin-resistant fragment from complexes of ribosomal protein S4 with 16-S RNA of Escherichia coli and from the uncomplexed protein; Newberry V et al.; A fragment of ribosomal protein S4 was prepared by limited trypsin degestion of a specific complex between protein S4 and 16-S RNA . It was characterised for amino acid sequence and the N-terminal 46 amino acids were found to be absent . An intermediate fragment, cut at Arg-43, was also observed at low trypsin concentrations . Evidence is presented that the protected fragment constitutes the primary RNA-binding region of the protein . No smaller protein fragments were found that rebound to the RNA . A mechanism for the degradation of the N-terminal region of the protein is proposed and two probable functions of the excised region are given . Under milder trypsin digestion conditions than for the complex, the same fragment, cut at Arg-46, was also prepared from the free protein . This result, together with that from a control experiment, indicates that at least within this local region, the protein conformation is conserved in both the free protein and the protein-RNA complex . This is the first direct evidence for the conservation of conformation in a protein when both complexed and uncomplexed with a ribosomal RNA.

Eur J Biochem, 1977 Jun 1, 76(1), 1 - 6
Subunit structure of the methionine-repressible aspartokinase II--homoserine dehydrogenase II from Escherichia coli K12; Dautry-Varsat A et al.; The quaternary structure of Escherichia coli K12 aspartokinase II--homoserine dehydrogenase II has been examined . This multifunctional protein has a molecular weight Mr = 176000 . It is composed of two subunits having the same molecular weight and the same charge . The results obtained from the examination of tryptic maps, the number and amino acid composition of cysteine-containing peptides and the uniqueness of the N-terminal sequence, strongly suggest that the 2 subunits are identical . The properties of aspartokinase II--homoserine dehydrogenase II can be compared to those of the much better known protein aspartokinase I--homoserine dehydrogenase I.

Chem Phys Lipids, 1977 Jun, 19(2), 99 - 106
Isosteres of natural phosphates . 6 . The preparation and properties of lysophosphotidic acid; Tang JC et al.; We herein report the first chemical synthesis of phosphonic acid analogues of lysophosphatidic acid . The racemic isosteric analogues, 4-acyloxy-3-hydroxybutyl-1-phosphonic acids, of lysophosphatidic acid were prepared by both catalytic and hydride reductions of the 4-acyloxy-3-oxobutyl-1-phosphonic acids, a general method for the preparation of the latter having been reported previously . The lysophosphatidic acids have been found to substrates for lysophosphatidic acid acyl transferase, and may be acylated chemically to yield phosphotidic acids . The latter reaction is of use in the preparations of differentially acylated phosphatidic acids.

Can J Biochem, 1977 Jun, 55(6), 618 - 24
The effect of proflavine on pyruvate kinase I of Escherichia coli B; McKellar RC et al.; Proflavine (PF) inhibited glucose use in sensitive but not resistant Escherichia coli B . Glucose transport (as measured by alpha-methylglucoside accumulation) was only partly inhibited by PF concentration that completely blocked glucose use . Fructose 1,6-diphosphate-(FDP)-regulated pyruvate kinase (PK1) (EC 2.7.1.40), the only glycolytic enzyme affected by PF, was completely inhibited by a dye concentration of 0.8 mM . The inhibition curve for PF was sigmoidal, suggesting that PF was acting as an allosteric inhibitor . PF increased the K 1/2 for phosphoenolpyruvate (PEP) and lowered the V; however, it had no effect on the Hill number for PEP . PF inhibition was partially reversed by FDP but not by cyclic AMP, AMP, ATP, fuctose 6-phosphate, or dithiothreitol . Studies with a variety of acridines indicated that those substituted at the 3-position are the most effective inhibitors and also that hydrophobic interactions may be involved in PF inhibition of PK I . PK I for E . coli B/Pr was also strongly inhibited by PF, indicating that PF resistance does not lie at the level of this enzyme . Ribose-5-phosphate-regulated pyruvate kinase (EC 2.7.1.40) was much less sensitive that PK I to the inhibitory effects of PF . A role for PF as a molecular probe for PK I has been proposed.

Biophys Chem, 1977 Jun, 7(1), 33 - 9
Association kinetics with coupled diffusion . An extension to coiled-chain macromolecules applied to the lac repressor-operator system; Berg OG et al.; The association of a molecule onto a specified binding site on a large chain-like macromolecule is described in the "sliding" model, where the molecule is allowed to move along the chain in a one-dimensional diffusion which is coupled to the three-dimensional diffusion in solution . The present work extends a previous one by treating the chains more generally as coiled instead of straight . The model is applied to the lac repressor-operator association . A general expression for the rate of unspecific attachment to a chain-like macromolecule is also derived.

Jpn J Surg, 1977 Jun, 7(2), 82 - 9
Occurrence of disseminated intravascular coagulation (DIC) in obstructive jaundice and its relation to biliary tract infection; Takeda S et al.; Coagulation studies were done on 78 consecutive cases of obstructive jaundice with or without biliary tract infection . Among 26 cases with biliary tract infection 20 cases showed no bleeding tendency but remarkable hypercoagulability with decreased fibrinolytic activity . Other six cases developed diffuse bleeding tendency in addition to the signs of hypotension and multiorgan dysfunction such as oliguria, respiratory distress and mental confusion . Most showed marked coagulation defects characterized by thrombocytopenia, decreased fibrinogen, antithrombin III and plasminogen levels and narrowing of maximal amplitude in thrombelastogram as well as the increase of fibrin degradation products and positive soluble fibrin monomer complexes . All except one died and three cases were autopsied . In two cases postmortem examination revealed multiple fibrin thrombi in lungs and other organs . A cause of the development of bleeding tendency in obstructive jaundice presently observed may likely to be due to the occurrence of disseminated intravascular coagulation (DIC), i.e . hypercoagulability caused by the biliary tract infection is responsible.

Ann Intern Med, 1977 Jun, 86(6), 714 - 8
Epidemic diarrhea at Crater Lake from enterotoxigenic Escherichia coli . A large waterborne outbreak; Rosenberg ML et al.; In June and July 1975, Gastrointestinal illness occurred in more than 200 staff members and 2000 visitors to an American national park . In was characterized by prolonged diarrhea, cramps, nausea, and vomiting, lasted a median duration of 8 days, and was significantly associated with consumption of park water (P less than 0.001), which had been contaminated by raw sewage . Enterotoxigenic Escherichia coli serotype 06:K15:H16 was isolated from 20 of 49 ill park residents and from the park's water supply, but not from 71 residents who had never been ill or had been well for at least 4 days . No other bacterial, viral, or parasitic pathogens were isolated from ill or well persons . This outbreak is the first waterborne epidemic of diarrheal illness shown to be due to enterotoxigenic . E . coli, and this study documents one mode of transmission of this pathogen . This investigation also suggests the relative insensitivity of current methods for identifying persons infected with this organism, either by the culturing of randomly selected isolates or by measuring serologic responses.

J Bacteriol, 1977 Jun, 130(3), 1393 - 6
Deoxyribonucleic acid strand breaks during freeze-drying and their repair in Escherichia coli; Ohnishi T et al.; Freeze-drying of Escherichia coli cells caused strand breaks of deoxyribonucleic acid (DNA) in both radiation-sensitive and -resistant strains . However, in the radiation-resistant strain E . coli B/r the damaged DNA was repaired after rehydration, whereas in the radiation-sensitive strain E . coli Bs-1 the damaged DNA was not repaired and the DNA was degraded . Repeated freeze-drying did not break the damaged DNA into smaller pieces.

J Bacteriol, 1977 Jun, 130(3), 1387 - 9
Inhibition of Histoplasma capsulatum ribonucleic acid polymerases by homologous and heterologous ribonucleic acid; McMillian R et al.; The ribonucleic acid (RNA) polymerases from the yeast phase of Histoplasma capsulatum are differentially sensitive to RNA isolated from the yeast and mycelial phases of this fungus and from Escherichia coli . Low-molecular-weight RNA from H . capsulatum was the most effective inhibitor.

J Bacteriol, 1977 Jun, 130(3), 1317 - 25
Identification of polypeptides necessary for chemotaxis in Escherichia coli; Silverman M et al.; Molecular cloning techniques were used to construct Escherichia coli-lambda hybrids that contained many of the genes necessary for flagellar rotation and chemotaxis . The properties of specific hybrids that carried the classical "cheA" and "cheB" loci were examined by genetic complementation and by measuring the capacity of the hybrids to direct the synthesis of specific polypeptides . The results of these tests with lambda hybrids and with a series of deletion mutations derived from the hybrids redefined the "cheA" and "cheB" regions . Six genes were resolved: cheA, cheW, cheX, cheB, cheY, and cheZ . They directed the synthesis of specific polypeptides with the following apparent molecular weights: cheA, 76,000 and 66,000; cheW, 12,000; cheX, 28,000; cheB, 38,000; cheY, 8,000; and cheZ, 24,000 . The presence of another gene, cheM, was inferred from the protein synthesis experiments . The cheM gene directed the synthesis of polypeptides with apparent molecular weights of 63,000, 61,000, and 60,000 . The synthesis of all of these polypeptides is regulated by the same mechanisms that regulate the synthesis of flagellar-related structural components.

J Bacteriol, 1977 Jun, 130(3), 1206 - 13
Determination of deoxyribonucleic acid replication time in exponentially growing Escherichia coli B/r; Churchward G et al.; The time necessary to replicate the chromosome (C period) was measured in Escherichia coli B/r (ATCC 12407) and a low-thymine-requiring derivative of that strain . In the Thy- strain, C was measured as a function of growth rate and exogenous thymine concentration either from step-up or chloramphenicol experiments . In the Thy+ parental strain, C was measured only as a function of the growth rate and only by the chloramphenicol method . The C period was found to decrease with growth rate and, in the Thy- strain, the C period also decreased with increasing thymine concentration . It approached a value of approximately 37 min at high growth rates.

J Bacteriol, 1977 Jun, 130(3), 1109 - 16
Rate of ribosomal ribonucleic acid chain elongation in Escherichia coli B/r during chloramphenicol treatment; Shen V et al.; In Escherichia coli B/r growing in glucose-amino acids medium, the radioactive labeling of 5S ribosomal ribonucleic acid (rRNA) and transfer RNA (tRNA) was measured after the simultaneous addition to the bacteria of chloramphenicol (CAM) (100 mug/ml), rifampin (200 mug/ml), and radioactive uracil . Accumulation of 5S rRNA ceased 85 s after the addition of rifampin, independent of the presence or absence of CAM; this indicates that CAM did not affect the rRNA chain growth rate . Together with previous measurements of the synthesis of rRNA and messenger RNA under these conditions, the results imply that CAM caused a redistribution of RNA polymerase which greatly favored stable RNA synthesis (77 to 97% of total functioning RNA polymerase engaged in synthesis of rRNA and tRNA) . Further, it is inferred that RNA polymerase molecules were activated that were inactive during exponential growth . The labeling of tRNA observed under these conditions suggests the existence of clusters of tRNA genes at the 3' end of long transcripts that resemble the rRNA precursor in length and response to CAM and may be parts of rRNA transcripts.

J Bacteriol, 1977 Jun, 130(3), 1072 - 83
Partial purification of glycerophosphate acyltransferase from Escherichia coli; Snider MD et al.; Glycerophosphate acyltransferase, a membrane-bound enzyme catalyzing the initial step of phospholipid biosynthesis in Escherichia coli, has been extracted with Triton X-100, a nonionic detergent, and purified 20- to 40-fold . This preparation is free from lysophosphatidate acyltransferase . Glycerophosphate acyltransferase is inactive in detergent extracts, but can be reconstituted by the addition of phospholipid . Under such conditions, the enzyme is associated with phospholipid . The sole product of the reaction with acyl coenzyme A as substrate is 1-acyl-sn-glycero-3-phosphate . Furthermore, the enzyme shows a marked preference for saturated fatty acyl conenzyme A, implying that this enzyme is responsible for the predominance of saturated moieties in position 1 of E . coli phospholipids . Acyltransferase from two mutants, plsA and plsB, was partially purified and characterized . Results support the view that plsB is a structural gene for the acyltransferase, but suggest that the plsA gene product is not directly involved in phospholipid biosynthesis.

J Bacteriol, 1977 Jun, 130(3), 1038 - 46
Menaquinone biosynthesis: mutants of Escherichia coli K-12 requiring 2-succinylbenzoate; Guest JR; Two independent mutants of Escherichia coli K-12, selected for their inability to grow anaerobically with fumarate as the terminal electron acceptor, were shown to be deficient in menaquinone biosynthesis . In both cases, exogenously supplied 2-succinylbenzoate promoted normal anaerobic growth on a lactate plus fumarate medium . Anaerobic growth of the mutants on glucose minimal medium was impaired but could be restored to normal by adding either uracil or 2-succinylbenzoate . The addition of 2-succinylbenzoate (but not uracil) permitted the synthesis of menaquinone and demethylmenaquinone by both mutants . The menaquinone content of the parental strain grown on lactate plus fumarate was three times greater than observed after growth on glucose . Transduction studies with phage P1 showed that the two mutations are very closely linked and probably affect the same gene, menC, which is cotransducible with nalA (23%), glpT (51%), and purF (8 to 14%) . The gene order nalA-nrdA-glpTA-menC-purF was indicated . The results were consistent with 2-succinylbenzoate being an intermediate in menaquinone biosynthesis and show that the gene designated menC (located at 48.65 min of the E . coli chromosome) is involved in the conversion of chorismate to 2-succinylbenzoate . It was also concluded that menaquinone is essential for electron transport to fumarate in E . coli.

J Bacteriol, 1977 Jun, 130(3), 1024 - 9
Regulation of amino acid transport in Escherichia coli by transcription termination factor rho; Quay SC et al.; Amino acid transport rates and amino acid binding proteins were examined in a strain containing the rho-120 mutation (formerly SuA), which has been shown to lower the rho-dependent, ribonucleic acid-activated adenosine triphosphatase activity to 9% of the rho activity in the isogenic wild-type strain . Tryptophan and proline transport, which occur by membrane-bound systems, were not altered . On the other hand, arginine, histidine, leucine, isoleucine, and valine transport were variably increased by a factor of 1.4 to 5.0 . Kinetics of leucine transport showed that the LIV (leucine, isoleucine, and valine)-I (binding protein-associated) transport system is increased 8.5-fold, whereas the LIV-II (membrane-bound) system is increased 1.5-fold in the rho mutant under leucine-limited growth conditions . The leucine binding protein is increased fourfold under the same growth conditions . The difference in leucine transport in these strains was greatest during leucine-limited growth; growth on complex media repressed both strains to the same transport activity . We propose that rho-dependent transcriptional termination is important for leucine-specific repression of branched-chain amino acid transport, although rho-independent regulation, presumably by a corepressor-aporepressor-type mechanism, must also occur.

Br J Clin Pharmacol, 1977 Jun, 4(3), 267 - 73
Pharmacokinetics of pivmecillinam; Parsons RL et al.; 1 The plasma concentration/time curves of ampicillin and mecillinam in normal subjects were measured after oral administration of ampicillin (500 mg) and pivmecillinam (400 and 600 mg) . 2 Similar plasma concentration/time curves of ampicillin and mecillinam in the starved normal subjects followed oral administration of ampicillin (500 mg) and pivmecillinam (600 mg) . 3 The plasma concentration/time curve of mecillinam was measured in the same normal subjects after oral administration of pivmecillinam (400 mg) with a reproducible standardized Lundh test meal . 4 There was no statistically significant difference in the plasma concentration/time curve of mecillinam after pivmecillinam/400 mg) and the meal compared with the plasma concentration/time curve after oral pivmecillinam (400 mg) was given to the same subjects when starved . After administration of pivmecillinam (400 mg) with meal, Tasc was significantly delayed beyond the value obtained when the subjects were starved . 5 The pharmacokinetics of pivmecillinam in coeliac disease are normal . This finding contrasts with previous studies on the pharmacokinetics of another pivaloyloxymethylpenicillin ester, pivampicillin, in this condition.

Nucleic Acids Res, 1977 Jun, 4(6), 2057 - 64
Escherichia coli DNA synthesis in vitro: insensitivity of ATP-dependent DNA repair to inhibition by novobiocin; Schneck PK et al.; Novobiocin, an effective inhibitor of DNA replicaion in Escherichia coli, is shown to have no effect on the ATP-dependent DNA repair carried out by toluenized cells after ultraviolet irradiation . Therefore novobiocin can be considered a selective inhibitor of replicative DNA synthesis in vitro.

Nucleic Acids Res, 1977 Jun, 4(6), 1803 - 13
Relaxed circular SV40 DNA as cleavage intermediate of two restriction endonucleases; Ruben G et al.; We have determined the mode of cleavage of superhelical SV40 DNA (Form I) by restriction endonucleases EcoRI and HpaII at 37 degrees C . By analysis with agarose gel electrophoresis and direct examination with dark field electron microscopy, we found that a large amount of the single-nicked circular DNA (Form II) was produced before the linear SV40 DNA (Form III) appeared . Thus, both restriction enzymes cleave only one strand of the superhelical DNA first . The second cleavage on the complementary strand occurred after a lag period . The first order rate constant for the second cleavage by EcoRI endonuclease was determined and a kinetic reaction scheme for both enzymes is proposed.

Proc Natl Acad Sci U S A, 1977 Jun, 74(6), 2446 - 50
Fidelity of synthesis of preribosomal RNA in isolated nucleoli and nucleolar chromatin; Ballal NR et al.; Comparisons were made of the T1 ribonuclease digests of 32P-labeled nucleolar 45S RNA of intact Novikoff hepatoma cells and the RNA synthesized in vitro by isolated nucleoli . Approximately 200 oligonucleotide spots were found in the two-dimensional chromatogram of 45S nucleolar RNA labeled in vivo, which includes fragments of 18S and 28S rRNA and nonconserved spacer regions; four spots containing 2'-O-methyl nucleotides were not found in the corresponding pattern of RNA labeled in vitro . This high degree of fidelity was retained in the patterns of spots from the RNA produced with nucleolar chromatin as template . This specific expression of rDNA was lost when the nucleolar chromatin was completely deproteinized . Specific spots found in the control patterns were absent and many nonspecific oligonucleotides were found to be labeled . A similar nonspecific chromatogram pattern was found when nucleolar chromatin was transcribed with RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) of Escherichia coli . These results show that specificity of genetic expression in vitro of isolated chromatin of eukaryotic systems is dependent on the chromatin-associated proteins and the type of RNA polymerase present.

Eur J Biochem, 1977 Jun 1, 76(1), 91 - 7
Analysis of the relA gene product of Escherichia coli; Pedersen FS et al.; The relA gene product, ATP: GTP 3'-pyrophosphotransferase (stringent factor) has been isolated in homogeneous form from an Escherichia coli strain polyploid for this gene at a yield of 1 mg/100 g cells and at a specific activity in a ribosome-activated assay at 37 degrees C of 120 mumol guanosine pentaphosphate formed min-1 mg protein-1 . The specific activity in a methanol-activated assay at 25 degrees C was found to be 4 mumol guanosine pentaphosphate formed min-1 mg protein-1 . These values are about 100 times higher than reported by others . Our further studies of this enzyme led to the following results . Antibodies raised against this enzyme inhibit the ribosome-activated synthesis of guanosine tetraphosphate and pentaphosphate but have no effect on the much slower synthesis, detected in the absence of ribosomes . The amount of stringent factor in the relA+ strain CP78 is estimated to about 1 copy per 200 ribosomes . The amount of antibody-binding material in CP79 (relA) is at least 5 times lower.

J Bacteriol, 1977 Jun, 130(3), 1399 - 401
Normal iron-enterochelin uptake in mutants lacking the colicin I outer membrane receptor protein of Escherichia coli; Soucek S et al.; The outer membranes of two independent colicin Ia-resistant mutants of Escherichia coli K-12 lack the colicin Ia receptor protein . Such mutants exhibit normal capacity for enterochelin (enterobactin)-mediated iron uptake . It is concluded that the colicin Ia receptor is not involved in iron-enterochelin uptake.

Br J Exp Pathol, 1977 Jun, 58(3), 255 - 9
Relationship between Escherichia coli endotoxin toxicity and the immunization status of normal adult guinea-pigs; McLean AJ; Lipopolysaccharide extracts of E . coli (endotoxins) were toxic to normal adult guinea-pigs, yet no evidence was found of serological reactivity to endotoxins or of "immediate" dermal reactivity in vivo . In contrast, testing of the serum of experimentally immunized animals provided evidence of precipitins, agglutinins and haemolysins . It is concluded that endotoxin exerts toxic effects in the absence of significant antibody production . This suggests that endotoxin toxicity is independent of immune processes not involving cell-mediated responses.

Pediatrics, 1977 Jun, 59(6), 827 - 32
Otitis media in children less than 12 weeks of age; Tetzlaff TR et al.; Cases of otitis media in infants under 12 weeks of age were reviewed to delineate the frequency, clinical features, and etiologic agents involved . Tympanocentesis was performed in 42 infants, 0 to 5 weeks of age, and in 17, from 6 to 11 weeks of age . The most common symptoms were irritability/lethargy (69%), fever (52%), cough (36%), vomiting (21%), diarrhea (20%), tachypnea (20%), and anorexia (18%) . Associated illnesses were present in 33 (54%) of the patients, the most common being pneumonia (9), bronchiolitis (7), meningitis (6), conjunctivitis (4), and omphalitis (4) . No peripartum infections or severe perinatal problems were found . Common respiratory pathogens were the predominant etiologic organisms, but coliform organisms were identified in 18% of the infants under 6 weeks of age . Cultures were sterile or grew organisms of questionable pathogenicity ("nonpathogens") in 39% of specimens . Since the signs and symptoms of otitis media in children less than 12 weeks of age are nonspecific and frequently associated with other major illnesses, the physician caring for these infants needs to be more aware of this disease and the therapeutic problems it presents.

Biochemistry, 1977 May 31, 16(11), 2384 - 9
Isolation and characterization of nonhistone chromosomal protein C-14 which stimulates RNA synthesis; James GT et al.; The nonhistone chromatin protein, C-14, was extracted from chromatin of Novikoff hepatoma ascites cells and isolated in high purity as shown by its migration as a single dense spot on two-dimensional polyacrylamide gels . Its mobility on sodium dodecyl sulfate gels is consistent with a molecular weight of approximately 70 000 . The amino acid composition shows that protein C-14 has an acidic:basic amino acid ratio of 1.8 . Its amino terminal amino acid is lysine . Protein C-14 stimulated the incorporation of {3H}UMP into RNA by approximately 30% when added to naked DNA and homologous RNA polymerase I . A 30% stimulation of {3H}UMP incorporation into RNA was also found when protein C-14 was added to an E . coli RNA polymerase system containing either E . coli or Novikoff hepatoma DNA.

J Biol Chem, 1977 May 25, 252(10), 3392 - 8
Use of 3H and 14C double-labeled glucose to assess in vivo pathways of amino acid biosynthesis in Escherichia coli; Csonka LN; 3H and 14C tracing data concerning amino acid biosynthetic pathways in Escherichia coli K12 are presented . Thirteen acidic and neutral amino acids were isolated from protein hydrolysates of wild type E . coli K12 grown aerobically or anaerobically in the presence of {U-14C}glucose together with {1-3H}glucose, {3-3H}glucose, {4-3H}glucose, or {6-3H}glucose . The observed 3H/14C counts of the amino acids were compared with the ratios expected on the basis of the input substrate specific activities and present understanding of biosynthetic pathways . For nine amino acids, serine, valine, leucine, threonine, isoleucine, glycine, glutamate, proline, and phenylalanine, the agreement between anticipated and observed specific activities was satisfactory . For the remaining four, methionine, alanine, aspartate, and (in cells labeled with {3-3H}glucose) tyrosine, the anticipated and observed specific activities differed markedly . For alanine, aspartate, and tyrosine, the differences are probably due to exchange of tritium in the course of biosynthesis; for methionine, it may be that there is a principle source of the methyl group other than carbon 3 of serine.

J Biol Chem, 1977 May 25, 252(10), 3538 - 47
Determination of the nucleotide sequence of part of the regulatory region for the galactose operon from Escherichia coli; Sklar J et al.; We have determined the sequence of 59 base pairs in the DNA preceding the site for initiation of transcription in the galactose operon . DNA from a lambdagal transducing phage was digested with restriction endonucleases to obtain a DNA fragment from the gal regulatory region . This fragment extends from 59 base pairs prior to the transcription initiation site through the 45 base pairs which specify the 5'-terminal sequence of gal mRNA . Analyses of RNA transcripts derived from this fragment and a variety of direct DNA sequence analyses allowed us to deduce the following sequence for the DNA in this fragment: (formula: see text) Position +1 corresponds to the site for initiation of gal mRNA synthesized in the presence of cyclic AMP and its receptor protein, CRP . Transcription experiments indicate, however, that this fragment lacks some element of the gal promoter required for the stimulation of transcription by CRP-cAMP . There are, nonetheless, some similarities between the gal sequence preceding the transcription start site and the sequences of other promoter regions . These include the heptamer sequence T-A-T-G-G-T-T (--12 to --8) and the sequence A-C-A-C-T-T-T (--36 to --30).

J Biol Chem, 1977 May 25, 252(10), 3446 - 58
Rabbit globin mRNA: analysis of T1 RNAse digestion fragments; Paddock GV et al.; Rabbit globin complementary DNA made with RNA-dependent DNA polymerase (reverse transcriptase) was used as a template for in vitro synthesis of 32P-labeled RNA and deoxysubstituted RNA . The sequences of the nucleotides in most of the fragments resulting from combined ribonuclease T1 and alkaline phosphatase digestion have been determined . In addition, the 3' nearest neighbor was determined for several fragments resulting from digestion with T1 ribonuclease . The utility of the deoxysubstitution technique was demonstrated by the ease with which the sequences of pyrimidine-rich fragments could be determined . Many sequences thus determined were long enough to fit uniquely with the alpha- or beta-globin amino acid sequences . The positions of these fits were found to be clustered, leading us to believe that only certain regions of the complementary DNA are transcribed by Escherichia coli RNA polymerase . Other unique characteristics of RNA synthesis from a complementary DNA template include a high yield of free poly(A) and the fact that one must use low rather than high salt buffers to obtain transcripts of high molecular weight.

J Biol Chem, 1977 May 25, 252(10), 3214 - 8
Isolation of glutamic acid methyl ester from an Escherichia coli membrane protein involved in chemotaxis; Kleene SJ et al.; We have isolated glutamic acid 5-methyl ester from an Escherichia coli protein that is involved in chemotaxis . The bacteria were first incubated with {methyl-3H}methionine under conditions which are known to result in methylation of the protein . The protein, isolated by gel electrophoresis, was then digested by successive treatment with three proteolytic enzymes . One of the products was {methyl-3H}glutamic acid 5-methyl ester, identified by comparison with an authentic sample in the following studies: (a) chromatography on an automatic amino acid analyzer, (b) chromatography on paper in two solvent systems, (c) chromatography on paper of the N-acetyl derivatives, and (d) stability of the ester bond to various pH conditions . No aspartic acid 4-methyl ester was found in the enzymatic digest . Treatment of the methylated protein with alkali released the radioactivity as {3H}methanol, which was identified by gas chromatography and by preparation of the 3,5-dinitrobenzoate.

J Biol Chem, 1977 May 25, 252(10), 3121 - 7
Phosphoenolypyruvate synthetase of Escherichia coli: molecular weight, subunit composition, and identification of phosphohistidine in phosphoenzyme intermediate; Narindrasorasak S et al.; Phosphoenolypyruvate synthetase of Escherichia coli has been shown to be a dimer of molecular weight 150,000 . The constituent subunits appear to be identical . The enzyme tends to dissociate to monomers at low protein concentration, but the tendency is much diminished in the phosphoenzyme form, suggesting that enzyme phosphorylation is accompanied by a structural rearrangement in the subunit contact domain . The enzyme appears to show half of the sites reactivity with respect to its phosphorylation by ATP . Several lines of evidence, including identification of 3-phosphohistidine in alkaline digests of the phosphoenzyme, indicate that a histidyl residue is the site of phosphorylation.

Biochim Biophys Acta, 1977 May 25, 487(2), 368 - 77
Specific phospholipid requirement for activity of the purified respiratory chain NADH dehydrogenase of Escherichia coli; Dancey GF et al.; The highly purified respiratory chain NADH dehydrogenase (EC 1.6.99.3) of Escherichia coli is inactive in the absence of detergent or phospholipid . Triton X-100 is the detergent that gives optimal activity, but the Triton X-100-activated enzyme is stimulated an additional 2-fold by E . coli phospholipids . Phosphatidylglycerol and diphosphatidylglycerol are the most effective lipid activators . The activated complex prepared with diphosphatidylglycerol is stable, whereas that with phosphatidylglycerol loses activity rapidly . Maximum activation by phospholipids occurs after preincubation at 0 degrees C and at pH 7 . Triton X-100 is required at low concentrations for lipid activation, but high concentrations interfere with the activation . When the enzyme is optimally activated by phospholipids, it may be additionally activated 2-fold by spermidine, but not by magnesium . In contrast, the Triton X-100-activated form of the enzyme is stimulated by several divalent cations, without specificity . Thus, the most stable, active form of the purified NADH dehydrogenase is generated in the presence of diphosphatidylglycerol and spermidine.

Mol Gen Genet, 1977 May 20, 153(1), 51 - 60
Transcription of insertion elements IS1 and IS2 in vitro; Besemer J et al.; Insertion elements IS1 and IS2 integrated within the gal operator-promoter region, an IS1 element in gene galT and insertions IS1 and IS2 integrated in the xycIIOP region of phage lambda were transcribed in vitro with E . coli RNA-polymerase . The insertion elements are transcribed exclusively by polymerase molecules started at the gal promoter and the lambdaPR promoter respectively . No promoter exists on IS1 or IS2 which can be recognized by RNA-polymerase in the pure in vitro transcription system used . Both insertions apparently are transcribed with a lower elongation rate than gal operon DNA or lambdaDNA . RNAs transcribed from the termini of IS1 and IS2 respectively were analysed by hybridization experiments . They are different in sequence.

Mol Gen Genet, 1977 May 20, 153(1), 45 - 9
Trans-dominance of dnaA mutants in Escherichia coli; Zahn G et al.; Temperature sensitivity of growth and DNA synthesis was tested in merogenotes heterozygous for the dnaA allele . All combinations tested (F dnaA+/dnaA5, F dnaA+/dnaA46, F dnaA+/dnaA204, F dnaA5/dnaA+, F dnaA204/dnaA+) were temperature sensitive . The mutant dnaA allele is thus trans-dominant to the wild type allele.

Mol Gen Genet, 1977 May 20, 153(1), 23 - 7
Dependence of DNA dark repair on protein synthesis in Escherichia coli; Sedliakova M et al.; We investigated the influence of amino-acidless treatments applied prior and after UV irradiation (AA-irradiated AA+; AA-irradiated A-; AA+ irradiated AA-) on survival, dimer excision, postirradiation DNA degradation, DNA synthesis and sedimentation profiles of parental DNA of E . coli B/r Hcr+ cells . In dependence on the treatment applied, the fluence 50 J/m2 yielded distinctly different fractions of survivors within 0,03-85% . In all cases dimers were completely excised . The rate of DNA degradation was similar during a 30-40 min period after UV during which the bulk of dimers was excised . Degradation ceased, however, earlier in the prestarved cells than in exponentially growing ones; it was prolonged by aminoacidless postincubation . Sedimentation profiles of parental DNA did not differ during the whole period of dimer excision . In AA+ AA- cells DNA synthesis was not restored for several hours after addition of amino acids . In AA- AA- cells addition of amino acids resulted in a fast resumption of DNA synthesis . We conclude that removal of dimers and repair of gaps were similar in all cases . We believe that aminoacidless treatments influence production and repair of damage to the sites of DNA replication . The treatment appears to prevent this damage when applied before UV irradiation, but interferes with its restoration when applied after UV irradiation . Consequently, the former treatment increases survival of cells while the latter produces an opposite effect.

Mol Gen Genet, 1977 May 20, 153(1), 1 - 4
Genetic analysis of mutations affecting ribonuclease II in Escherichia coli; Ono M et al.; Exonuclease activity in an Escherichia coli K12 mutant S296 is less than 1% of that in the wild type strain (Nikolaev et al., 1976) . Another mutant N464 has thermolabile ribonuclease II (Castles and Singer, 1968; Kuwano et al., 1969) . Genetic analysis of these mutants by Hfr conjugation and P1 transduction indicates that the structural gene (rnb) for ribonuclease II is located near the pyrF gene (28 min on the E . coli genetic map of Bachmann, Low and Taylor (1976)), and the most probable gene order is tyrT-trp-pyrF-rnb.

Biochim Biophys Acta, 1977 May 17, 476(2), 97 - 107
Induction and repair of strand breaks and 3'-hydroxy terminals in the DNA of mammalian cells in culture following gamma-ray irradiation; Matsudaira H et al.; DNA was isolated in a fairly pure and intact state from cultured mouse leukaemia cells (L5178Y) after gamma-ray irradiation using a hydroxyapatite column chromatography method, and analysed further by sucrose gradient centrifugation or DNA polymerase (EC 2.7.7.7, enzyme A of Klenow from Escherichia coli) assay . Irradiation of the cells induced single- and double-strand breaks in the DNA with an efficiency of 100 eV/break and 1300 eV/break, respecitvely . Approximately 50% of the single-strand breaks were estimated to be those arising from allali-labile lesions . A linear, dose-dependent increase was found in the template activity of the DNA, indicating the induction of 3'-OH terminals by gamma-irradiation . Post-irradiation incubation of the cells in serum-free medium allowed the majority of the breaks to rejoin within a few hours . Repair of the alkali-labile lesions was, however, found to be much slower than that of "actual" single-strand breaks . A slight increase of the DNA template activity was found during the period of post-irradiation incubation . The reason for the increase is discussed.

Biochemistry, 1977 May 17, 16(10), 2213 - 20
Modification-deficient transfer ribonucleic acids from relaxed control Escherichia coli: structures of the major undermodified phenylalanine and leucine transfer RNAs produced during leucine starvation; Kitchingman GR et al.; The structures of the major, chromatographically unique phenylalanine and leucine tRNAs produced during leucine starvation of a relaxed control (rel-) mutant of E . coli have been determined . The results demonstrate that the unique species are modification-deficient forms of the major, normally occurring isoacceptor species . The unique tRNAphe differs from the fully modified species at nucleotide positions 16, 37, 39, 47, and 55 from the 5' terminus . The unique species contains uridine (U) in place of dihydrouridine-16 (D16), isopentenyladenosine in place of 2-thiomethyl-N6-(delta2-isopentenyl)adenosine-37, a mixture of U and pseudouridine (psi) in position 39, a mixture of U and 3-(3-carboxypropyl)uridine at position 47, and a mixture of U and psi at position 55 . The chromatographically normal isoacceptor from amino acid starved cells is deficient in D16 and psi55, indicating that that species is a mixture of mature and undermodified tRNAs . The unique tRNALeu isoacceptor consists of two subspecies which are undermodified forms of the major, normally occurring isoacceptor, tRNALeuI . Both unique subspecies lack the D and psi residues which occur at positions 16 and 39 from the 5' terminus; one subspecies also lacks D17 . Compared with the tRNALeusI from wild-type strains of E . coli B and K12, both tRNALeuI from nonstarved cells and the unique, rel-tRNALeu are deficient in the modified guanosine which normally occurs adjacent to the anticodon and the pseudouridine in the GTpsiC sequence of the psi loop . Both the unique tRNAPhe and the unique tRNALeu lack dihydrouridine residues which occur in the 5' half of the D loop and pseudouridines which occur in the 3' half of the anticodon loop and adjoining stem . Taken together, these findings suggest that the same enzymes are responsible for the formation of these particular modified bases in both tRNAs . The results further suggest that several, perhaps most, of the tRNAs from cells cultured under conditions in which RNA and protein synthesis are uncoupled will be similarly deficient in dihydrouridine and pseudouridine and other minor nucleosides which occur less frequently . Because both modification-deficient rel-tRNAs have dihydrouridine at position 20 and pseudouridine in the psi loop (and at position 41 in the unique tRNALeu), the results support the view that there was multiple D-and psi-forming enzymes in E . coli, some of which may turn over rapidly or are selectively inactivated when protein synthesis is blocked . The results are discussed with a view toward understanding the structural basis for the altered biological activity of the unique tRNAPhe species and the order of events in the posttranscriptional modification of newly synthesized tRNA.

Biochemistry, 1977 May 17, 16(10), 2095 - 100
High-resolution nuclear magnetic resonance determination of transfer RNA tertiary base pairs in solution . 2 . Species containing a large variable loop; Hurd RE et al.; The number of base pairs in the solution structure of several class III D3VN tRNA species from E . coli has been determined by analyzing the number of low-field (-15 to -11 ppm) proton resonances in their nuclear magnetic resonance spectra at 360 MHz . Contrary to previous reports indicating the absence of tertiary resonances, all the spectra exhibit the expected number of secondary base pair resonances plus approximately ten extra resonances derived from tertiary base pairs in the three-dimensional folding of these molecules . The possible origins of some of these tertiary resonances are discussed; none of the spectra exhibits the characteristic resonance of the 8-14 tertiary base pair seen in class I D4V5 tRNA spectra.

Biochemistry, 1977 May 17, 16(10), 2086 - 94
High-resolution nuclear magnetic resonance determination of transfer RNA tertiary base pairs in solution . 1 . Species containing a small variable loop; Reid BR et al.; Eight class I tRNA species have been purified to homogeneity and their proton nuclear magnetic resonance (NMR) spectra in the low-field region (-11 to -15 ppm) have been studied at 360 MHz . The low-field spectra contain only one low-field resonance from each base pair (the ring NH hydrogen bond) and hence directly monitor the number of long-lived secondary and tertiary base pairs in solution . The tRNA species were chosen on the basis of their sequence homology with yeast phenylalanine tRNA in the regions which form tertiary base pairs in the crystal structure of this tRNA . All of the spectra show 26 or 27 low-field resonances approximately 7 of which are derived from tertiary base pairs . These results are contrary to previous claims that the NMR spectra indicate the presence of resonances from secondary base pairs only, as well as more recent claims of only 1-3 tertiary resonances, but are in good agreement with the number of tertiary base pairs expected in solution based on the crystal structure . The tertiary base pair resonances are stable up to at least 46 degrees C . Removal of magnesium ions causes structural changes in the tRNA but does not result in the loss of any secondary or tertiary base pairs.

Biochemistry, 1977 May 17, 16(10), 1064 - 73
Proton nuclear magnetic resonance study of the effect of pH on tRNA structure; Steinmetz-Kayne M et al.; The low-field 220-MHz proton nuclear magnetic resonance (NMR) spectra of four tRNA molecules, Escherichia coli tRNAPhe, tRNA1Val, and tRNAfMet1, and yeast tRNAPhe, at neutral and mildly acidic pH are compared . We find a net increase in the number of resonances contributing to the -9.9-ppm peak (downfield from sodium 4,4-dimethyl-4-silapentanesulfonate) in three of these tRNAs at pH 6, while tRNAfMet1 does not clearly exhibit this behavior . The increase in intensity at this resonance position is half-completed at pH 6.2 in the case of yeast tRNAPhe . An alteration at the 5'-phosphate terminus is not involved, since removal of the terminal phosphate does not affect the gain in intensity at -9.9 ppm . Based on a survey of the tertiary interactions in the four molecules, assuming that they possess tertiary structures like that of yeast tRNAPhe at neutral pH, we tentatively attribute this altered resonance in E . coli and yeast tRNAPhe to the protonation of the N3 of the adenine residue at position 9 which results in the stabilization of the tertiary triple A23-U12-A9 . This intepretation is supported by model studies on the lowfield proton NMR spectrum of AN oligomers at acid pH, which reveal an exchanging proton resonance at -9.4 ppm if the chain length N greater than or equal to 6.

Tijdschr Diergeneeskd, 1977 May 15, 102(10), 619 - 29
{Cross-infection during scalding and plucking of broiler chickens (author's transl)}; Mulder RW et al.; In two poultry-processing plants, experiments were carried out to detect cross-infection during scalding and plucking of broiler chickens . These trials showed that cross-infection was only detectable when experimental infection with the indicator micro-organism Escherichia coli K12 was applied externally to the broiler chickens . When infection was applied internally, only a small degree of transmission was observed . Indicator micro-organisms applied externally to broiler chickens prior to scalding and plucking, were present on the cooled product in larger numbers than those applied internally in the intestines of broilers . The number of carcasses which were positive after cooling was found to have decreased in poultry-processing plant B compared with the situation after plucking, whereas this number was not affected to any appreciable extent in processing plant A.

Biochim Biophys Acta, 1977 May 12, 482(1), 52 - 63
Purification and molecular properties of the AMP-activated pyruvate kinase from Escherichia coli; Somani BL et al.; The AMP-activated pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) from Escherichia coli has been purified 200 times through a three-step procedure which gives a homogeneous preparation with a specific activity of 110 . The enzyme appears to be a tetramer of molecular weight 190 000 . Subunits (molecular weight 51 000) show a single amino-terminal amino acid (serine) and appear as a single band in polyacrylamide gel electrophoresis in sodium dodecyl sulphate . The enzyme crystallizes in conditions of reduced dielectric constant of the solvent in the pH range 6.5-7.5 . Kinetic and regulatory properties of the purified enzyme are similar to those described for crude preparations of the enzyme.

Biochim Biophys Acta, 1977 May 12, 482(1), 11 - 8
Lipoamide dehyrogenase immobilized on porous glass; Scouten WH et al.; Lipoamide dehydrogenase (NADH:lipoamide oxidoreductase, EC 1.6.4.3) isolate from pig heart and Escherichia coli was covalently coupled by both diazonium and amide bonds to controlled pore glass beads (96% silica) . When the enzyme was immobilized in the presence of NAD+, the enzyme no longer exhibited its normal requirement for NAD+ for full activity . If the immobilized enzyme was then treated with NADase, the requirement for NAD+ was restored . Enzyme immobilized in the absence of NAD+ exhibited normal NAD+ dependence both prior to an after NADase treatment . These results are discussed in terms of co-immobilization of NAD+ at or near the allosteric site of the enzyme.

J Biol Chem, 1977 May 10, 252(9), 3102 - 4
Immunospecific retention of oligonucleotides possessing N6-methyladenosine and 7-methylguanosine; Munns TW et al.; Antibodies specific for N6-methyladenosine (m6A) and for 7-methylguanosine (m7G) were immobilized on Sepharose and the resulting immunoadsorbents tested for their ability to retain specific oligonucleotides possessing the corresponding antigenic haptens (i.e . m6A and m7G) . Results obtained with oligonucleotides derived from ribonuclease T1 digests of Escherichia coli tRNA (previously labeled with {methyl-3H}methionine) indicated that each immunoadsorbent quantitatively and exclusively retained those methyl-3H-labeled oligonucleotides possessing {methyl-3H}m6A and {methyl-3H}m7G . Elution and subsequent characterization of the retained methyl-3H-labeled oligonucleotides via DEAE-cellulose chromatography revealed the presence of several small oligonucleotides containing m7G and a single, larger oligonucleotide containing m6A . These findings are in accord with previously sequenced structures which indicate that numerous bacterial tRNA species possess m7G while only tRNAVal contains m6A.

J Biol Chem, 1977 May 10, 252(9), 3064 - 73
Multiple pathways for primary processing of ribosomal RNA in Escherichia coli; Gegenheimer P et al.; A comparison of isogenic RNase III+ and RNase III- strains of Escherichia coli shows that although both synthesize precursor and mature 16 S and 23 S ribosomal RNAs, the transient rRNA species of the RNase III- strain differ from those of the RNase III+ strain . The RNase III+ strain synthesizes p16 and p23 rRNA, whereas the RNase III- strain produces unstable 17 S, 18 S, "p23," 25 S and 30 S RNA molecules . The 30 S RNA, which is a primary transcript of the ribosomal RNA gene cluster, does not contribute significantly to any of the smaller RNAs, nor is m23 rRNA derived from 25 S but rather from "p23" RNA . Mature 16 S rRNA is derived from both 18 S and 17 S RNA, and 17 S RNA can be derived from 18 S . Additionally, an unstable RNA species about 300 bases long is missing in the RNase III- strain and another species which seems to be about 50 bases larger appears . Processing of the primary ribosomal RNA transcript in RNase III- strains of Escherichia coli is accomplished during its transcription by two independent pathways which are not so utilized in RNase III+ strains . One pathway yields 18 S and precursor 23 S RNAs which are processed to mature rRNAs; the second pathway yields 25 S RNA and perhaps 16 S rRNA . The second pathway, unlike the first, is inhibited by chloramphenicol treatment . At slow rates of ribosomal RNA synthesis, the nascent transcript is processed preferentially by the first pathway . We suggest that in the absence of RNase III, which is involved in the primary processing of rRNA in E . coli, other enzymes involved in primary and secondary processing of rRNA in RNase III+ cells can recognize their sites on the nascent rRNA transcript and accomplish the primary processing.

J Biol Chem, 1977 May 10, 252(9), 2881 - 90
An aspartate transcarbamylase lacking catalytic subunit interactions . Study of conformational changes by ultraviolet absorbance and circular dichroism spectroscopy; Kerbiriou D et al.; A modified form of aspartate transcarbamylase is synthesized by Escherichia coli in the presence of 2-thiouracil which does not exhibit homotropic cooperative interactions between active sites yet retains heterotropic cooperative interactions due to nucleotide binding . The conformational changes induced in the modified enzyme by the binding of different ligands (substrates, substrate analogs, a transition state analog, and nucleotide effectors) were studied using ultraviolet absorbance and circular dichroism difference spectroscopy . Comparison of the results for the modified enzyme and its isolated subunits to those for the native enzyme and its isolated subunits showed that the conformational changes detected by these methods are qualitatively similar in the two enzymes . Comparison of the absorbance difference spectra due to the binding of a transition substrate analog to the intact native or modified enzymes to the corresponding results for the isolated subunits suggested that ligand binding causes an increased exposure to solvent of certain tyrosyl and phenylalanyl residues in the intact enzymes but not in the isolated subunits . This result is consistent with a diminution of subunit contacts due to substrate binding in the course of homotropic interactions in the native enzyme . Such conformational changes, though perhaps necessary for homotropic cooperativity, are not sufficient to cause homotropic cooperativity since the modified enzyme gave identical perturbations . Interactions of the transition state analog, N-(phosphonacetyl)-L-aspartate, with the modified enzyme were studied . Enzyme kinetic data obtained at low aspartate concentrations showed that this transition state analog does not stimulate activity, but rather exhibits the inhibition predicted for the total absence of homotropic cooperative interactions in the modified enzyme . Spectrophotometric titrations of the number of catalytic sites with the transition state analog showed that the modified enzyme and its isolated subunits possess, respectively, four and two high affinity sites for the inhibitor instead of six and three observed in the case of the normal enzyme and its isolated catalytic subunits . These results are correlated with the lower specific enzymatic activities of the modified enzyme and its catalytic subunits compared to the normal corresponding enzymatic species.

J Biol Chem, 1977 May 10, 252(9), 2873 - 80
Functionally important arginine residues of aspartate transcarbamylase; Kantrowitz ER et al.; The reaction of phenylglyoxal with aspartate transcarbamylase and its isolated catalytic subunit results in complete loss of enzymatic activity (Kantrowitz, E . R., and Lipscomb, W . N . (1976) J . Biol . Chem . 251, 2688-2695) . If N-(phosphonacetyl)-L-aspartate is used to protect the active site, we find that phenylglyoxal causes destruction of the enzyme's susceptibility to activation by ATP and inhibition by CTP . Furthermore, CTP only minimally protects the regulatory site from reaction with this reagent . The modified enzyme still binds CTP although with reduced affinity . After reaction with phenylglyoxal, the native enzyme shows reduced cooperativity . The hybrid with modified regulatory subunits and native catalytic subunits exhibits slight heterotropic or homotropic properties, while the reverse hybrid, with modified catalytic subunits and native regulatory subunits, shows much reduced homotropic properties but practically normal heterotropic interactions . The decrease in the ability of CTP to inhibit the enzyme correlates with the loss of 2 arginine residues/regulatory chain (Mr = 17,000) . Under these reaction conditions, 1 arginine residue is also modified on each catalytic chain (Mr = 33,000) . Reaction rate studies of p-hydroxymercuribenzoate, with the liganded and unliganded modified enzyme suggest that the reaction with phenylglyoxal locks the enzyme into the liganded conformation . The conformational state of the regulatory subunit is implicated as having a critical role in the expression of the enzyme's heterotropic and homotropic properties.

J Biol Chem, 1977 May 10, 252(9), 2808 - 14
A new endonuclease from Escherichia coli acting at apurinic sites in DNA; Ljungquist S; A new DNA endonuclease has been purified 3000-fold from Escherichia coli . The enzyme specifically catalyzes the formation of single strand breaks at apurinic and apyrimidinic sites in DNA, but has no activity on intact or single-stranded DNA . Further, the enzyme shows little or no activity on heavily ultraviolet-irradiated DNA, but cleaves x-irradiated DNA, presumably at apurinic and apyrimidinic sites introduced by the radiation treatment . The enzyme, which is tentatively named endonuclease IV, has no detectable associated exonuclease or DNA N-glycosidase activity and does not seem to be identical with any previously known E . coli endonuclease . Endonuclease IV has no Mg2+ requirement, and is fully active in the presence of EDTA . Enzyme activity is stimulated by 0.2 to 0.3 M NaCl and is unusually salt-resistant . Further, the enzyme is fairly heat-stable, and is not inhibited by tRNA . The sidimentation coefficient, S(o)20,w, is 3.4 S . It seems that endonuclease IV is active in DNA repair.

J Biol Chem, 1977 May 10, 252(9), 2802 - 7
Endonuclease from Escherichia coli that acts specifically upon duplex DNA damaged by ultraviolet light, osmium tetroxide, acid, or x-rays; Gates FT et al.; An endonuclease which is active upon DNA exposed to ultraviolet light at a photoproduct other than thymine dimers has been extensively purified from Escherichia coli . The small (2.7 S) enzyme is active in the presence of EDTA, has a neutral pH optimum, and is inhibited by tRNA and 1 M NaCl . It has no detectable exonuclease, DNA-N-glycosidase, or ribonuclease activities . The enzyme also nicks duplex DNA exposed to OsO4, x-rays, or acid, but it does not act upon undamaged DNA or irradiated single-stranded DNA . The majority of sites of action in DNA exposed to ultraviolet light or OsO4 appear to be alkali-stable, but those in DNA exposed to x-rays or acid are not . The incisions created by the endonuclease contain 5'-phosphate termini . The enzyme is possibly the same as E . coli endonuclease III described by Radman (Radman, M . (1976) J . Biol . Chem . 251, 1438-1445), but it is distinguishable from the other endodeoxyribonucleases described from that organism.

Biochemistry, 1977 May 3, 16(9), 1988 - 96
Effects of a trinucleotide ethyl phosphotriester, Gmp(Et)Gmp(Et)U, on mammalian cells in culture; Miller PS et al.; The nonionic 2'-O-methyribooligonucleotide ethyl phosphotriester, Gmp(Et)Gmp(Et)U, is complementary to the...ApCpC...sequence found in the amino acid accepting stem of most tRNAs and the anticodon region of tRNAgly and to the threonine codon of mRNA . Gmp(Et)Gmp(EtU forms hydrogen-bonded complexes with the amino acid accepting stem of tRNApheyeast and unfractionated tRNA Escherichia coli under physiological salt conditions at 37 degrees C as determined by equilibrium dialysis . The extent of phenylalanine aminoacylation of tRNApheE.coli is inhibited 39% by Gmp(Et)Gmp(Et)U at 37 degrees C in solution . The triester is resistant to hydrolysis by serum nucleases and cell lysates . The triester is readily taken up by transformed Syrian hamster fibroblasts growing in monolayer . Within the cell, the triester is deethylated to give the trinucleotide species Gmp(Et)GmpU, GmpGmp(Et)U, and GmpGmpU and is also hydrolyzed to dimeric and monomeric units . Treatment of transformed fibroblasts in monolayer with 25 micronM Gmp(Et)Gmp(Et)U results in a 40% inhibition of cellular protein synthesis with a concurrent slight increase in cellular RNA synthesis during the first 4 h . After 4 h, the rate of cellular protein synthesis begins to recover while RNA synthesis returns to that of the control . Our biochemical studies suggest that inhibition of cellular protein synthesis might be expected if Gmp(Et)Gmp(Et)UGmp(Et)GmpU, GmpGmp(Et)U, and GmpGmpU, which have been taken up by or formed within the cell, physically bind to tRNA and mRNA and inhibit the function of these nucleic acids . The reversible inhibition of protein synthesis may be a consequence of further degradation of the trinucleotide species within the cell as well as to an increase in supply of RNA molecules involved in protein synthesis . The growth of the transformed fibroblasts is inhibited during the first 24 h of incubation with 25 micronM Gmp(Et)Gmp(Et)U after which growth proceeds at a normal rate . In cloning experiments, the number and size of colonies formed by the transformed fibroblasts after 5 days exposure to 25 micronM triester is decreased by 50% relative to untreated controls . The temporary inhibition of cell growth may reflect the transitory inhibition of cellular protein synthesis caused by the triester.

Biochim Biophys Acta, 1977 May 3, 476(1), 32 - 7
Discrimination between bromouracil and thymine for uptake into DNA in drm- and dra- mutants of Escherichia coli K12; Ryderg B; The relative efficiency of bromouracil and thymine for uptake into DNA was measured in various thymine-requiring strains of Escherichia coli K12 . It was found that: 1 . Mutants with genotype thyA- dra- discriminate against bromouracil to much greater extent than do mutants with genotype thyA- drm- . 2 . The discrimination in dra-mutants is dependent on thymine concentration, whereas discrimination in drm- mutants is almost independent of thymine concentration . It is suggested that the intracellular level of deoxyribose 5-phosphate affects the efficiency of uptake into DNA of bromouracil relative to thymine.

Biochemistry, 1977 May 3, 16(9), 1814 - 9
Stimulation, by two Escherichia coli supernatant proteins, of the initiation of polypeptide synthesis; Tsuda Y et al.; Two protein factors (A and B) have been partially purified from Escherichia coli supernatant which, in combination, are more effective than 0.5 M NH4Cl in stimulating ribosomes for AcPhe-tRNA and fMet-tRNA binding, for the puromycin reaction, and for incorporating acetylphenylalanine from AcPhe-tRNA into polypeptide . The factors appear to differ from the initiation factors, the elongation factor EF-T, and ribosomal proteins . Some uncertainty exists as to whether factor B is different from EF-G . To maximize the effect of the factors in initiator tRNA binding, we preincubated the ribosomes with the factors and carried out the binding assay for a short period at 15 degrees C . Maximal stimulation of binding occurred after about a 2-min preincubation at 37 degrees C . Longer preincubation times were required at 15 degrees C, and only slight stimulation was observed after preincubation at 0 degrees C . The extent of stimulation by the factors was not affected when the NH4Cl concentration was increased from 40 to 500 mM in the preincubation . The presence of both the 30S and 50S ribosomal subunits is required for the enhancement of AcPhe-tRNA binding . Polyphenylalanine synthesis carried out without AcPhe-tRNA is inhibited by the factors . It is suggested that the factors may act by inducing a structural rearrangement of the ribosomes.

Biochemistry, 1977 May 3, 16(9), 1795 - 801
In vitro transcription of Moloney leukemia virus genes in infected cell nuclei and chromatin: elongation of chromatin associated ribonucleic acid by Escherichia coli ribonucleic acid polymerase; Shih TY et al.; The in vitro transcription of viral specific DNA sequences in nuclei and chromatin isolated from mouse cells chronically infected with Moloney murine leukemia virus (Mo-MuLV) has been studied . The in vitro RNA synthesized by Escherichia coli RNA polymerase has been isolated by sulfhydryl affinity column following reaction in the presence of 5-mercuriuridine triphosphate . By comparison of the Crt curves of the in vitro RNA with that of 70S viral RNA, the content of viral sequences is found to be 1.3% in nuclei product and 0.24% in chromatin product which is lower than the 2.5% found in chromatin associated RNA . This latter value, however, is very close to the in vivo viral RNA content in pulse-labeled {3H}RNA of the infected cells . Unexpectedly, it is observed that over 20% of the chromatin associated RNA prelabeled in vivo with {5-3H}uridine is elongated and tagged with Hg atoms during RNA synthesis catalyzed by the exogenous E . coli RNA polymerase in the presence of Hg-UTP . The elongation reaction is dependent on the presence of all four nucleotide triphosphates and appears to be due to E . coli RNA polymerase per se . It is suggested that most of the viral specific sequences observed in the in vitro RNA products are very likely initiated and derived from the chromatin associated species . The implication of the present findings for in vitro RNA synthesis in nuclei and chromatin as related to regulation of gene expression is discussed.

Biochemistry, 1977 May 3, 16(9), 1765 - 72
Studies on gene control regions . 1 . Chemical synthesis of lactose operator deoxyribonucleic acid segments; Goeddel DV et al.; The chemical synthesis of lactose operator DNA segments is described . The 31-base-paired duplex contains the DNA recognized by lac repressor protein and twofold rotationally symmetric base pairs on either side of the tight binding region . The synthesis includes the deoxyoligonucleotides d(T-G-T-G-G), d(A-A-T-T-G-T-G-A-G), d(C-G-G-A-T-A-A-C-A-A-T-T), d(T-C-A-C-A), d(T-G-T-G-A-A-A-T-T-G-T), d(T-A-T-C-C-G-C-T-C-A-C), and d(A-A-T-T-C-C-A-C-A) . These deoxyoligonucleotides were characterized by two-dimensional sequencing techniques, paper chromatography, and thin-layer chromatography.

Biochemistry, 1977 May 3, 16(9), 1890 - 6
Fluorescent derivatives of the pyruvate dehydrogenase component of the Escherichia coli pyruvate dehydrogenase complex; Papadakis N et al.; One sulfhydryl group per polypeptide chain of the pyruvate dehydrogenase component of the pyruvate dehydrogenase multienzyme complex from Escherichia coli was selectively labeled with N-{P-(2-benzoxazoyl)phenyl}-maleimide (NBM), 4-dimethylamino-4-magnitude of-maleimidostilbene (NSM), and N-(4-dimethylamino-3,5-dinitrophenyl)maleimide (DDPM) in 0.05 M potassium phosphate (pH 7) . Modification of the sulfhydryl group did not alter the enzymatic activity or the binding of 8-anilino-1-naphthalenesulfonate (ANS) or thiochrome diphosphate to the enzyme . The fluorescence of the NBM or NSM coupled to the sulfhydryl group on the enzyme was quenched by binding to the enzyme of the substrate pyruvate the coenzyme thiamine diphosphate, the coenzyme analogue thiochrome diphosphate, the regulatory ligands acetyl-CoA, GTP, and phosphoenolpyruvate, and the acetyl-CoA analogue, ANS . Fluorescence energy transfer measurements were carried out for the enzyme-bound donor-acceptor pairs NBM-ANS, NBM-thiochrome diphosphate ANS-DDPM, and thiochrome diphosphate-DDM . The results indicate that the modified sulfhydryl group is more than 40 A from the active site and approximately 49 A from the acetyl-CoA regulatory site . Thus, a conformational change must accompany the binding of ligands to the regulatory and catalytic sites . Anisotropy depolarization measurements with ANS bound on the isolated pyruvate dehydrogenase in 0.05 M potassium phosphate (pH 7.0) suggest that under these conditions the enzyme is dimeric.

Eur J Biochem, 1977 May 2, 75(1), 43 - 53
Synthesis of exported proteins by membrane-bound polysomes from Escherichia coli; Randall LL et al.; A membrane-bound fraction of polysomes of Escherichia coli has been isolated after lysis of cells without the use of lysozyme . Protein-synthesis studies in vitro show that membrane-bound and free polysomes are different in the following respects . 1 . Membrane-bound polysomes synthesize proteins which are exported from the cell . The products include proteins of the outer membrane and a secreted periplasmic protein, the maltose-binding protein . 2 . The major product synthesized by free polysomes is elongation factor Tu, a soluble cytoplasmic protein . 3 . The activity of membrane-bound polysomes in vitro is more resistant to puromycin than is the activity of free polysomes . In addition, the mRNA associated with membrane-bound polysomes is more stable than the bulk of cellular mRNA as revealed by studies with rifampicin.

Biochim Biophys Acta, 1977 May 2, 466(3), 393 - 401
Phase transitions of phospholipid bilayers from an unsaturated fatty acid auxotroph of Escherichia coli; Uehara K et al.; Total phospholipids were extracted from cells of temperature sensitive unsaturated fatty acid auxotrophs of Escherichia coli (K-12 UFAts) grown at 28degrees C (PL28), and at 42degrees C in the presence of 2% KCl as an osmotic stabilizer (PL42 (KCl)) . From the analysis of fatty acids, it was shown that the content of unsaturated fatty acids of PL42 (KCl) is only 9% of the total fatty acids, while that of PL28 is 54% . The thermal phase transitions of the bilayers prepared from the phospholipid fractions were studied by proton magnetic resonance . The line widths of the methylene signals and the sums of the methylene and methyl signal intensities were plotted against reciprocal values of absolute temperature 1/T or temperature itself . From the plots phase transitions were detected at about 19degrees C for PL28 and at 43degrees C for PL42 (KCl) . In spite of its complex composition of fatty acids a highly cooperative transition was observed in the case of PL42 (KCl) . It was also suggested that the phospholipids bilayers in the biomembranes of this strain at the growth temperature (42 degrees C) are in the state where the gel and liquid crystalline phases coexist.

Eur J Biochem, 1977 May 2, 75(1), 217 - 24
Uridine phosphorylase from Escherichia coli . Physical and chemical characterization; Leer JC et al.; Uridine phosphorylase from Escherichia coli has been purified to homogeneity . The enzyme was found to have a molecular weight of 176000 and to consist of 8 probably identical subunits with molecular weights of 22000 . These numbers were determined from equilibrium centrifugations in the analytical ultracentrifuge, from dodecylsulphate gel electrophoresis and from amino acid analysis . Moreover the following physico-chemical constants were determined: s020,w = 8.2 x 10(-13) s, upsilon2 = 0.751 cm3/g, A1%280 (1 cm) = 6.73 and a specific activity of 183 units/mg towards uridine . The enzyme shows some activity towards deoxyuridine and thymidine . The activity is not impaired through substitution by bromo, fluoro or methyl groups in the 5-position of the uracil base, but no enzymatic activity is observed when cytosine base is used in the nucleoside substrate.

Res Vet Sci, 1977 May, 22(3), 267 - 70
The effect of Escherichia coli endotoxin on plasma histaminase activity in the domestic fowl and the involvement of caeruloplasmin; Butler EJ et al.; The injection of four-to nine-week-old fowls with Escherichia coli O111:B4 endotoxin (0-1 and 1-0 mg/kg) produced a two to eight fold rise in the histaminase activity of the plasma 24 h afterwards . In some cases this increase was still detectable after 48 h . This activity was strongly correlated with the p-phenylenediamine oxidase activity of caeruloplasmin . Electrophoretic studies with 7-5 per cent polyacrylamide gels indicated that fowl caeruloplasmin also was histaminase and putrescinase activity and that the release of this protein from the liver by endotoxin is largely responsible for the increase in the activity of the plasma . In untreated fowls this activity was lower than published values for several mammals and does not explain the relative resistance of the fowl to the acute effects of endotoxins and large doses of histamine.

Infect Immun, 1977 May, 16(2), 617 - 22
Assay of Escherichia coli heat-labile enterotoxin with vero cells; Speirs JI et al.; The continuous cell line of African green monkey kidney, Vero, showed characteristic morphological changes in response to culture filtrates from toxigenic strains of Escherichia coli . The response compared favorably with that of Y-1 (mouse adrenal) and CHO (Chinese hamster ovary) cells . Vero cells were the simplest and most economical to maintain in the laboratory.

Br J Cancer, 1977 May, 35(5), 595 - 601
A test for mutation theory of cancer: carcinogenesis by misrepair of DNA damaged by 4-nitroquinoline 1-oxide; Kondo S; Evidence for a mutation theory of cancer is presented by reviewing the experimental work on 4-nitroquinoline 1-oxide (4NQO) carcinogenesis . 4NQO almost completely mimics u.v . light and produces 4NQO-purine adducts on DNA . When 4NQO-treated cells are held in liquid medium under appropriate conditions, the 4NQO adducts disappear from DNA, in parallel to decrease of premutational damage in Escherichia coli, or pretransformational damage in cultured mouse cells . Post-treatment with caffeine greatly diminishes the yields by 4NQO of mutants in E . coli, malignant transformants in cultured mouse cells and tumour nodules in the lung of mice . Potentially tumourigenized stem cells in the lung remain sensitive to selective killing by caffeine for at least 5 days after 4NQO treatment, in spite of their DNA being apparently replicated, an indication that carcinogen-damaged DNA in the stem cell can be transmitted to its successive daughter stem cells for many generations . This peculiar characteristic is discussed as a possible lead to the crux of the mutation theory of cancer in vivo, and a model for carcinogenesis is proposed.

J Bacteriol, 1977 May, 130(2), 968 - 71
Transport of 3,4-dihydroxybutyl-1-phosphonate, an analogue of sn-glycerol 3-phosphate; Leifer Z et al.; 3,4-Dihydroxybutyl-1-phosphonate (DHBP), an analogue of glycerol 3-phosphate, is actively transported by the sn-glycerol 3-phosphate transport system of Escherichia coli strain 8 . The Km for the transport of DHBP is 200 microM.

J Bacteriol, 1977 May, 130(2), 960 - 2
Nucleoside triphosphate pools in minicells of Escherichia coli; Manwaring JD et al.; The nucleoside triphosphate pools of Escherichia coli minicells are different from those in parental cells . The growth phase in which minicells accumulate significantly affects the pool sizes.

J Bacteriol, 1977 May, 130(2), 957 - 9
Coordinate control of the synthesis of ribonucleoside diphosphate reductase components in Escherichia coli; Fuchs JA; Ribonucleoside diphosphate reductase subunits B1 and B2 and ether-permeabilized cell activities of Escherichia coli increase in parallel during thymine deprivation . Thioredoxin and thioredoxin reductase activities are not affected by thymine deprivation.

J Bacteriol, 1977 May, 130(2), 951 - 3
Multivalent regulation of isoleucine-valine transaminase in an Escherichia coli K-12 ilvA deletion strain; Kline EL et al.; In a strain of Escherichia coli K-12 lacking threonine deaminase, the enzyme converting alpha-ketoisovalerate and alpha-keto-beta-methylvalerate to valine and isoleucine, respectively, was multivalently repressed by valine, isoleucine, and leucine . This activity was due to transaminase B, specified by the ilvE structural gene.

J Bacteriol, 1977 May, 130(2), 911 - 36
Polysaccharide capsule of Escherichia coli: microscope study of its size, structure, and sites of synthesis; Bayer ME et al.; This report describes the structure, size, and shape of the uncollapsed polysaccharide capsule of Escherichia coli strain Bi 161/42 {O9:K29(A):H-}, its ultrastructural preservation as well as the filamentous components of the isolated capsular material . In a temperature-sensitive mutant, sites were localized at which capsular polysaccharide is "exported" to the cell surface . The highly hydrated capsule of the wild-type cells was visible in the uncollapsed state after freeze-etching, whereas dehydration in greater than or equal to 50% acetone or alcohol caused the capsule to collapse into thick bundles . This was prevented by pretreatment of the cell with capsule-specific immunoglobulin G; the capsule appeared as a homogeneous layer of 250- to 300-nm thickness . The structural preservation depended on the concentration of the anti-capsular immunoglobulin G . Temperature-sensitive mutants, unable to produce capsular antigen at elevated temperatures, showed, 10 to 15 min after shift down to permissive temperature, polysaccharide strands with K29 specificity appearing at the cell surface at roughly 20 sites per cell; concomitantly, capsule-directed antibody started to agglutinate the bacteria . The sites at which the new antigen emerged were found in random distribution over the entire surface of the organism . Spreading of purified polysaccharide was achieved on air-water interfaces; after subsequent shadow casting with heavy metal, filamentous elements were observed with a smallest class of filaments measuring 250 nm in length and 3 to 6 nm in width . At one end these fibers revealed a knoblike structure of about 10-nm diameter . The slimelike polysaccharides from mutants produced filamentous bundles of greater than 100-microns length, with antigenic and phage-receptor properties indistinguishable from those of the wild-type K29 capsule antigen.

J Bacteriol, 1977 May, 130(2), 900 - 10
Area of 16S ribonucleic acid at or near the interface between 30S and 50S ribosomes of Escherichia coli; Santer M et al.; To determine the region of 16S ribonucleic acid (RNA) at the interface between 30 and 50S ribosomes of Escherichia coli, 30 and 70S ribosomes were treated with T1 ribonuclease (RNase) . The accessibility of 16S RNA in the 5' half of the molecule is the same in 30 and 70S ribosomes . The interaction with 50S ribosomes decreases the sensitivity to T1 RNase of an area in the middle of 16S RNA . A large area near the 3' end of 16S RNA is completely protected in 70S ribosomes . The RNA near the 3' end of the molecule and an area of RNA in the middle of the molecule appear to be at the interface between 30 and 50S ribosomes . One site in 16S RNA, 13 to 15 nucleotides from the 3' end, normally inaccessible to T1 RNase in 30S ribosomes, becomes accessible to T1 RNase in 70S ribosomes . This indicates a conformational change at the 3' end of 16S RNA when 30S ribosomes are associated with 50S ribosomes.

J Bacteriol, 1977 May, 130(2), 787 - 92
Flagellar formation in Escherichia coli electron transport mutants; Bar Tana J et al.; Mutants of Escherichia coli lacking ubiquinone or heme have been tested for motility and found to be essentially immotile . The loss of motility is identified with the loss of flagellum synthesis.

J Bacteriol, 1977 May, 130(2), 781 - 6
Escherichia coli K-12 tolF mutants: alterations in protein composition of the outer membrane; Chai TJ et al.; Outer membrane materials prepared from three independently isolated spontaneous Escherichia coli tolF mutants contained no detectable protein Ia . The loss of this protein was nearly completely compensated for by an increase in other major outer membrane proteins, Ib and II . Thus, the major outer membrane proteins accounted for 40% of the total cell envelope protein in both tol+ and tolF strains . No changes were found in the levels of inner membrane proteins prepared from tolF strains when compared with similar preparations from the tol+ strain . Phage-resistant mutants were selected starting with a tolF strain by using either phage TuIb or phage PA2 . These phage-resistant tolF strains contained neither protein Ia nor protein Ib . The mutation leading to the loss of protein Ib in these strains is independent of the tolF mutation and is located near malP on the E . coli genetic map.

J Bacteriol, 1977 May, 130(2), 775 - 80
Iodination of Escherichia coli with chloramine T: selective labeling of the outer membrane lipoprotein; Munford RS et al.; Iodination of Escherichia coli cells with chloramine T preferentially labels the free and murein-bound forms of the outer membrane lipoprotein . Iodination for 15 s at 15 degrees C labels the two forms of the lipoprotein almost exclusively, whereas iodination for 60 s at 25 degrees C also labels the other major outer membrane proteins . Chloramine T iodination is a rapid, simple technique for labeling the outer membrane lipoprotein.

J Bacteriol, 1977 May, 130(2), 724 - 8
Cell division in Escherichia coli BS-12 is hypersensitive to deoxyribonucleic acid damage by ultraviolet light; Bridges BA et al.; Escherichia coli BS-12 uvrA lon is hypersensitive to ultraviolet light . On minimal agar plates at densities in excess of about 10(7) bacteria per plate, as few as one or two photoreversible pyrimidine dimers in the entire genome are sufficient to cause inhibition of cell division . Most of the resulting filaments are unable to divide or form a viable colony . Inhibition of cell division appears to be a rapid consequence of replication of deoxyribonucleic acid containing a pyrimidine dimer . Photoreversibility of the inhibition of cell division persists indefinitely, indicating that the continued presence of the pyrimidine dimers (or the continued generation of daughter strand gaps) is necessary to maintain the division-inhibited state . In view of the kinetics for the production of filamentation by ultraviolet light and the extremely low average inducing fluence (0.03 J/m2), it is concluded that the initiating signal is not the same as that causing other inducible phenomena such as prophage induction or Weigle reactivation.

J Bacteriol, 1977 May, 130(2), 692 - 7
Deoxyribonucleic acid synthesis after inhibition of initiation of rounds of replication in Escherichia coli B/r; Bremer H et al.; The theory describing the effect of inhibition of initiation of rounds of deoxyribonucleic acid (DNA) replication on the accumulation of DNA is derived, and an analysis is presented which allows the determination of the time C taken to replicate the bacterial chromosome from the kinetic changes in the accumulation of DNA . This analysis is applied to experiments in which inhibition of initiation was achieved by inhibiting protein or protein and ribonucleic acid synthesis with chloramphenicol or rifampin . The results for both antibiotics are identical and indicate that there is a delay of 6 to 11 min in the effect of the antibiotics on initiation of rounds of replication . If this delay is taken into account, then the value of the C period estimated from such experiments agrees with values obtained by other methods, whereas by conventional data evaluation of such experiments the C period would be overestimated . In the low thymine-requiring derivative of Escherichia coli B/r ATCC 12407 used here, the C period was found to be between 38 and 41 min for cultures growing with a mass doubling time of 29 min in glucose-amino acids medium, supplemented with 20 micrograms of thymine/ml.

J Bacteriol, 1977 May, 130(2), 684 - 91
Mutations in the L-arabinose operon of Escherichia coli B/r with reduced initiator function; Gonzalez IL et al.; Partial reversion mutants derived from a strain containing a strongly polar initiator-defective mutation (araI1036) in the L-arabinose operon were found to have several characteristics expected of mutants with reduced initiator function . These reversion mutations are cotransduced with the ara region and are probably within the araI region . Furthermore, they permit induction of the L-arabinose operon to a level only one-third of the normal wild-type level . These partially functional initiator regions reduce the expression of structural genes in the cis position only; they function quite independently of wild-type or defective initiator regions in the trans position . These mutants exhibit a two- to threefold increase in the rate of expression of ara operon genes within one-tenth of a generation after a shift of the growth temperature from 28 to 42 degrees C . This suggests that the temperature optimum for initiation of operon expression is higher for the partial revertant strains than it is for strains containing a wild-type initiator region.

J Bacteriol, 1977 May, 130(2), 667 - 75
Deoxyribonucleic acid repair in Escherichia coli mutants deficient in the 5'----3' exonuclease activity of deoxyribonucleic acid polymerase I and exonuclease VII; Chase JW et al.; A series of Escherichia coli strains deficient in the 5'----3' exonuclease activity associated with deoxyribonucleic acid (DNA) polymerase I (exonuclease VI) and exonuclease VII has been constructed . Both of these enzymes are capable of pyrimidine dimer excision in vitro . These strains were examined for conditional lethality, sensitivity to ultraviolet (UV) and X-irradiation, postirradiation DNA degradation, and ability to excise pyrimidine dimers . It was found that strains deficient in both exonuclease VI (polAex-) and exonuclease VII (xseA-) are significantly reduced in their ability to survive incubation at elevated temperature (43 degrees C) beyond the reduction previously observed for the polAex single mutants . The UV and X-ray sensitivity of the exonuclease VI-deficient strains was not increased by the addition of the xseA7 mutation . Mutants deficient in both enzymes are about as efficient as wild-type strains at excising dimers produced by up to 40 J/m2 UV . At higher doses strains containing only polAex- mutations show reduced ability to excise dimers; however, the interpretation of dimer excision data at these doses is complicated by extreme postirradiation DNA degradation in these strains . The additional deficiency in the polAex xseA7 double-mutant strains has no significant effect on either postirradiation DNA degradation or the apparent deficiency in dimer excision at high UV doses observed in polAex single mutants.

J Bacteriol, 1977 May, 130(2), 656 - 60
Genetic analysis of Escherichia coli O111:B4, a strain of medical and biochemical interest; Coleman WG Jr et al.; Procedures have been worked out which allow, for the first time, the genetic analysis of Escherichia coli O111:K58:H2 (O111:B4) . The approximate map position of mutant loci was determined by mating with 15 Hfr strains of E . coli K-12 . In addition, P1 transduction procedures were used for establishing relative gene order and linkage for any region of the E . coli O111:B4 chromosome . To obtain these, it was necessary to select for a rare P1 lysogen since E . coli O111:B4 is resistant to phage P1 . Finally, genetic homology between E . coli strains K-12 and O111:B4 is suggested since they can form stable haploid hybrids, and several loci have similar map positions in the two strains.

J Bacteriol, 1977 May, 130(2), 642 - 55
In vitro transcription of the Escherichia coli K-12 argA, argE, and argCBH operons; Sens D et al.; Deoxyribonucleic acid isolated from argA and argECBH transducing phages was utilized to study the in vitro synthesis of argA, argE, and argCBH messenger ribonucleic acid . The specific regulation of these operons by the arginine holorepressor was demonstrated, providing evidence that the majority, if not all, of the control of these operons is exercised at the transcriptional level . Data are presented which indicate that the arginine holorepressor functions by binding to the operator region and concomitantly prevents the binding of ribonucleic acid polymerase to the corresponding promoter region.

Mol Biol (Mosk), 1977 May-Jun, 11(3), 656 - 60
{Influence of protein bound with single-stranded DNA on the synthesis of RNA and poly(A) . III . Protein product of F1 gene 5}; Polonskii IuS et al.; Influence of the protein product of F1 phage gene 5 (protein 5) on the synthesis RNA and poly(A) in vitro was studied . It has been shown, that protein 5 has no effect on the transcription of the native DNA by E . coli RNA-polymerase, but completely prevents RNA and poly(A) synthesis on the denatured or single-stranded DNA at the protein/DNA ratio 10:1 . Protein 5 inhibits poly(A) synthesis with oligo(dT)9 and oligo(dT)12 as a template, preventing binding of enzyme to the oligonucleotide . After the initiation of the poly(A) synthesis the inhibition becomes considerably wearer . The biological function of the inhibition of the transcription by "unwinding" proteins is discussed.

Mol Biol (Mosk), 1977 May-Jun, 11(3), 611 - 9
{Repression of the enzyme inducible syntheses in Escherichia coli K12 mutant with a deleted ptsH gene}; Gershanovich VN et al.; The genome of lambda phage with thermosensitive repressor was integrated into the pts region of the E . coli chromosome . Such a lysogenic culture behaves as a pts mutant at 30 degrees . Heating of cells of this strain leads to the induction of lambda prophage and formation of deletions in the pts region . A mutant with a deletion covering ptsH gene was isolated after prophage induction . The deletion nature of pts mutation was confirmed in genetic and biochemical experiments . It was shown that the deletion is small and does not involve ptsI and lig genes . The isolated deltaptsH mutant possesses all characteristics of pts mutants: pleiotropic impairment of transport and utilization of a number of carbohydrates, repression of the enzyme inducible synthesis and resistance to catabolite repression with glucose . These data (together with earlier ones) allow us to conclude that the phosphorylated form of HPr is involved (in direct of indirect manner/ in activation of DNA transcription.

Mol Biol (Mosk), 1977 May-Jun, 11(3), 598 - 610
{Addition of the fluorescent label to the 3'-OH end of DNA and the 3'-OH end of nascent RNA}; Rozovskaia TA et al.; 3'(2')-O-acyl derivatives of the uridine triphosphate were synthesized . Acyl residues contained fluorescent dye; fluoresceine or rodamine C . Optical properties and stability of UTP analogues were studied . Their ability to serve as the substrates for calf thymus terminal deoxyribonucleotidyl transferase and E . coli RNA polymerase was also examined . It was shown that both enzymes were able to use tested analogues as substrates . Incorporation of the analogues into nascent RNA and DNA chains inhibited the synthetic reaction because of primer inactivation . The rate of the incorporation of the analogues showed an exponential time dependence

Mol Biol (Mosk), 1977 May-Jun, 11(3), 498 - 506
{Induction and repair of breaks in DNA chains in vivo due to the imbalance between DNA and protein synthesis}; Fradkin GE et al.; A sudden induction of the imbalance between the rates of DNA and protein synthesis in the cell (by nalidixic acid or by thymine starvation) results in the stabilization of breaks in DNA chains in vivo . Such "imbalance induced breaks" represent gaps in DNA chains formed with the participation of exonuclease V . Stabilization of the "imbalance induced breaks" is accompanied by DNA degradation and cell death . Restoration of the imbalance between the rates of DNA and protein syntheses by balanced inhibition completely prevents the stabilization of breaks in cells with competent repair systems . Balanced inhibition of intracellular DNA and protein synthesis decreases the rate of repair and permits to see the sequence of induction (stabilization) and disappearance (repair) of breaks in DNA chains in vivo . Repair of breaks occuring on the background of balanced inhition of DNA and protein synthesis decreases the extent of DNA degradation and completely prevents death of E . coli cells with completely functional repair systems.

Gene, 1977 May, 1(3-4), 255 - 80
In vitro packaging of a lambda Dam vector containing EcoRI DNA fragments of Escherichia coli and phage P1; Sternberg N et al.; In this report we describe a coliphage lambda vector system for cloning endo R . EcoRI DNA fragments . This system differs significantly from those previously described in two ways . First, restricted and ligated DNA is encapsidated in vitro . Second, with increasing lambda DNA size in the range 78 to 100% that of wild-type, the efficiency of DNA encapsidation into infectious phage particles markedly increases . For lambda wild-type DNA the efficiency of in vitro packaging (10(6) to 10(7) plaques produced per microgram of added DNA) is equal to, or better than, the standard CaCl2 transfection method . The use of a Dam mutation to facilitate recognition of size classes of inserted fragments is described . Using this vector and in vitro packaging, several E . coli and phage P1 and R.EcoRI fragments were cloned.

Gene, 1977 May, 1(3-4), 229 - 39
Cloning and amplification of alpha and beta mouse globin gene sequences synthesised in vitro; Rougeon F et al.; New chimeric Escherichia coli plasmids containing alpha or beta globin gene sequences of the mouse were constructed . Double-stranded DNA, synthesised in vitro in a 2-step reaction from mouse globin mRNA was inserted into E . coli plasmid pCR1, after tailing of the 2 DNAs with dG and dC respectively . Some of the mouse globin plasmids described contain at least 90% of the globin mRNA sequence and therefore contain the entire translated sequence of the globin genes . Some possible uses of these recombinant plasmids are described.

Gene, 1977 May, 1(3-4), 185 - 207
Sequence organization and expression of a yeast plasmid DNA; Gubbins EJ et al.; Saccharomyces cerevisiae strain A364A D5 contains circular double-stranded DNA molecules of 6230 +/- 30 base pairs (2mu DNA) which are present in 68 copies per cell and make up 2.4% of the haploid genome . About 0.4% of non-poly A containing yeast RNA hybridizes to the yeast DNA circles . When denatured and then self-annealed, the DNA molecules assume a characteristic "dumbbell" shape in the electron microscope indicating that each circle possesses a non-tandem inverted repeat sequence of 630 +/- 10 base pairs . Eco-RI digestion of purified 2mu DNA yields 4 fragments on an agarose gel whose combined molecular mass is twice that of the monomer circle, suggesting that there are 2 populations of circles, each of the same molecular weight . Representatives of each population have been separated by cloning in Escherichia coli via the bacterial plasmid pSC101 . Heteroduplex analysis of the cloned circles show that the 2 different populations arise because of intramolecular recombination between the inverted repeat sequences . Acrylamide gel patterns of polypeptides synthesized in bacterial mini-cells containing the hybrid plasmids between 2mu DNA and pSC101 are significantly different than the pattern obtained from mini-cells containing pSC101 alone.

Vopr Virusol, 1977 May-Jun, (3), 271 - 4
{Splitting and rejoining simian adenovirus type 7 DNA by using Eco RI restriction endonuclease and DNA-ligase}; Kislina OS et al.; Optimal conditions for ligation of simian adenovirus type 7 (SA-7) DNA fragments formed under the effect of treatment of the intact molecule with Eco RI endonuclease were established . It was shown that up to 30% of the original material may be ligated and transferred into a structure with molecular weight 23 X 10(6) daltons which corresponded to the molecular weight of the intact SA-7 DNA . The data of the existence of one recognition site for Eco RI in SA-7 DNA were confirmed . The biological activity of the ligated material was demonstrated on lambda-III phage DNA.

J Biochem (Tokyo), 1977 May, 81(5), 1525 - 30
Strain specificity of outer membrane proteins in Escherichia coli; Ichihara S et al.; Outer membrane proteins of various strains of Escherichia coli were compared using three different systems of sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis . The outer membranes of E . coli K-12, E . coli B, and E . coli J-5 had distinctive protein compositions . As regards proteins which interact with peptidoglycan, E . coli K-12 contained O-8 and O-9, while E . coli B possessed one protein which migrated to the position of O-9 . Although E . coli J-5 possessed two such proteins, O-8' and O-9', their positions on polyacrylamide gel were different from those of O-8 and O-9 . Protein O-7, which migrates slightly more slowly than O-8, was found specifically in E . coli K-12 . Proteins O-10 and O-11 were found in all strains tested, although the relative amounts were different depending on the strain . Strains of E . coli K-12 and E . coli J-5 gave three major bands, O-2a, O-2b, and O-3, in the region of high molecular weight . These proteins were repressed by iron in the cultivation media . Strains of E . coli B, on the other hand, gave only O-2b and O-3 . E . coli J-5 gave two other major bands in this region, but the amounts were not controlled by iron in the cultivation media.

J Biochem (Tokyo), 1977 May, 81(5), 1357 - 65
Properties of proteins produced after damage to deoxyribonucleic acid of Escherichia coli; Yoda K et al.; Large amounts of extra proteins, X (in the envelope fraction) and X' (in the cytoplasmic fraction) were detected by SDS-polyacrylamide gel electrophoresis when DNA of Escherichia coli was damaged . These two proteins had the same apparent molecular weight (appproximately 40,000) and were produced under identical conditions, including requirement for the recA" and lexA+ genotype . Sucrose density gradient centrifugation revealed that protein X' consisted of relatively large and heterogeneous aggregates in the cytoplasmic fraction; the distribution of protein X in the envelope was not determined . As proteins X and X' were shown to be equivalent, it is suggested that they are identical . Co-precipitation of the aggregates of protein X' with the envelope led to the appearance of protein X in the envelope fraction.

Biokhimiia, 1977 May, 42(5), 828 - 32
{Some peculiarities of phage DDVI-specific methylases}; Nikol'skaia II et al.; The types of methylases are found in the cellular extract of Escherichia coli B, infected with phage DDVI . One of them is a cellular enzyme, which methylates adenine to form 6-methylaminopurine (6-MAP) and is repressed in the infected cell in vivo . The second type, which is not found in the non-infected cells, is specific for phage DDVI and induces the formation of 7-methylguanine (7-MG) . Both enzymes recognize various sites, which accounts for the ratio 6-MAP/7-MG to vary in heterological DNAs between 2.07 in phage Sd DNA and 0.40 in phage DDII DNA . During in vitro incubation with homologous methylases phage DDVI DNA and especially phage T2 DNA are subjected to further methylation, which is probably indicative of their "undermethylation" in vivo . The DDVI-specific enzyme, similar to B-specific type, methylates DNA with a normal set of nitrogenous bases (phages Sd and DDII), as well as DNAs containing 5-oxymethylcytosine and glucose (phages T2 and DDVI) . Both methylases under study use only native double-helical DNA as substrate and are strongly inhibited by S-adenosylhomocysteine . Phage DDVI Methylase is characterized by low stability.

J Protozool, 1977 May, 24(2), 294 - 6
Correlation of encystment and division in Schizopyrenus russelli; Rastogi AK et al.; Schizopyrenus russelli, a free-living soil ameba, grows and encysts in the presence of bacteria . The encystment occurs with decline in the division rate . This is accompanied by incorporation of {U-14C} glucose into cyst cellulose . The degree of multiplication (but not of encystment) is a function of bacterial concentration . Berenil, a trypanocidal drug, while allowing excystment, completely inhibited multiplication of emerged amebae and their encystment . Addition of this drug after 24 hr, when amebae had gone into a phase of active division failed to check encystment, although it still inhibited further multiplication of the amebae . The findings suggest that a phase of cell division may be a prerequisite for encystment.

Appl Environ Microbiol, 1977 May, 33(5), 1233 - 6
Changes in lipid composition of Escherichia coli resulting from growth with organic solvents and with food additives; Ingram LO; Cells of Escherichia coli contain an altered fatty acid and phospholipid composition when grown in the presence of sublethal concentrations of a variety of organic solvents and food additives . The diversity of compounds examined which caused these changes indicates that no single catabolic pathway is involved . Many of the observed changes are consistent with the hypothesis that cells adapt their membrane lipids to compensate for the presence of these compounds in the environment . Both sodium benzoate and calcium propionate caused the synthesis of unusual fatty acids.

Appl Environ Microbiol, 1977 May, 33(5), 1207 - 8
Osmoregulation in symbiosis-independent mutants of Bdellovibrio bacteriovorus; Varon M et al.; Bdellovibrios capable of axenic growth grow in a cell-free medium at a rate considerably lower than that attainable in a two-membered culture with Escherichia coli . The axenic growth rate may be improved either by adjustment of the osmosity of the medium or by the addition of low concentrations of spermine.

Antibiotiki, 1977 May, 22(5), 422 - 6
{Comparative study of the physicochemical properties of the surface of Escherichia, their sensitivity to ampicillin and tetracycline and their capacity to absorb tetracycline}; Popova NA et al.; Development of resistance to ampicillin and tetracycline in Escherichia resulted in an increase in the electrokinetic potential and a decrease in the level of hydration and isoelectric values of pH . The changes in the hydration level mainly depended on the accompanying dissociation . Studies on 3H-tetracycline binding revealed a low accumulation capacity of the resistant mutants . The rate of 3H-tetracycline binding did not depend on the changes in the physico-chemical parameters of the cell surface due to resistance and dissociation.

Int J Radiat Biol Relat Stud Phys Chem Med, 1977 May, 31(5), 451 - 8
The effect of U.V.-irradiation on lambda DNA transcription; Ranade SS; The effect of U.V.-irradiation of template DNA has been studied in vitro in the E . coli RNA polymerase system with native and U.V.-treated lambda DNA . Lambda DNA is more susceptible to U.V . than is calf-thymus DNA, yet a residual activity is observed at a U.V . dose of 0-5+10(4) erg/mm2 . From the kinetic analysis of the reaction and the incorporation of lambda 32P-labelled nucleoside triphosphates, it seems reasonable to conclude that U.V.-irradiation probably does not affect the DNA initiation sites, recognizable by RNA polymerase . The transcription products made with U.V.-irradiated lambda DNA are assymmetrical, and hybridized to the right half (R) and the left half (L) of lambda DNA with the ratio of R/L = 4/1, and they show a lower hybridizability than the transcripts with native lambda DNA . The initiation sites recognizable by RNA polymerase seem to be the same on both native and U.V.-irradiated lambda DNA though the transcription of U.V.-treated lambda DNA appears to terminate with rather short RNA chains.

Int J Radiat Biol Relat Stud Phys Chem Med, 1977 May, 31(5), 407 - 13
Action of hydrogen peroxide on degradation of DNA after irradiation in Escherichia coli; Keller KM et al.; Hydrogen peroxide (H2O2), which produces breaks in cellular DNA, has not hitherto been shown to cause degradation of DNA . In this investigation it is shown that if transcription is blocked with rifampin, treatment with H2O2 causes degradation of DNA to nearly the same extent as does gamma-radiation . Further, if cells are given a treatment with H2O2 and incubated for 50 min, the amount of degradation in a second treatment is markedly less . This is attributed to the induction of the inhibitor of post-irradiation degradation of DNA (prd) by the first treatment . There is thus a double action of H2O2: first, to induce inhibition, and second, to cause degradation of DNA to begin in non-induced cells . The genetic dependence of induction by H2O2 mimics that of ionizing radiation . Accordingly, the induction process does not occur in recA- and lex- cells, because they are not inducible and is absent in recB- cells because they lack exonuclease V, the major component of prd . Potassium iodide (KI), an OH radical scavenger, negates the action of peroxide on DNA . The results obtained in this study suggest a possible theory for the evolution of radiation response systems

Gut, 1977 May, 18(5), 351 - 5
Escherichia coli serotypes throughout the gastrointestinal tract of patients with intestinal disorders; Tabaqchali S et al.; The O and H serotypes of Escherichia coli that were present along the entire length of the gastrointestinal tract of patients with small intestinal bacterial overgrowth were studied . Multiple sero- and biotypes were represented, although usually a single serotype predominated in each patient . In a number of cases the different O:H serotypes were antigenically related indicating that antigenic degradation was occurring . The serotypes isolated from the stomach and small intestine were represented in the faeces . In general, within the limitations of this study, there appears to be a stable ecosystem in each patient and it may require specific oral antibiotics to alter it.

Eur J Immunol, 1977 May, 7(5), 257 - 63
Ontogeny of the antibody-forming cell line in mice . III . The generation of mature anti-sheep red blood cell-specific B cells is antigen-dependent; Rosenberg YJ et al.; A role for antigen in the generation of fully mature splenic type B cells has been shown . In adoptive transfer experiments, cells from bone marrow or fetal liver required a longer period to give an anti-sheep red blood cell plaque-forming cell (PFC) response than those from spleen . This delay was not overcome by allowing the cells a 7-day sojourn in the irradiated host before antigen challenge . A two-stage protocol was designed in which the in vivo generation of fully mature cells could be measured by their ability to give PFC in lipopolysaccharide-stimulated cultures in vitro . These experiments showed that a critical factor which influences the final differentiation of bone marrow or fetal liver cells into mature, splenic type B cells is exposure to antigen.

Cell, 1977 May, 11(1), 181 - 5
Anomalous synthesis of ppGpp in growing cells; Gallant J et al.; In E . coli cells, accumulation of ppGpp is normally triggered by conditions that restrict the aminoacylation of tRNA or interfere with carbon/energy source metabolism; in both cases, the nucleotide's accumulation is associated with control of stable RNA synthesis and is generally believed to bring it about . We have found an anomalous situation wherein vigorously growing cells accumulate a high level of ppGpp and there is no restriction of stable RNA synthesis . This occurs when wild-type cells are shifted up from an abnormally low growth temperature to one in the optimal range (35 degrees C-40 degrees C) . The effect is partly, but not entirely, dependent upon the presence of a functional relA gene product . These results appear to call into question the simpler interpretations of the role of ppGpp in the control of stable RNA synthesis.

Proc Natl Acad Sci U S A, 1977 May, 74(5), 1865 - 9
Origin of replication of colicin E1 plasmid DNA; Tomizawa JI et al.; Cleavage maps of colicin E1 plasmid DNA and its smaller derivative, pNT1 DNA, were constructed by using restriction endonucleases . The nucleotide sequence of a region that contains the orgin of replication was determined . The site of the nucleotide from which DNA replication is initiated was determined with 6S L-fragments, the DNA fragment first made on colicin E1 plasmid DNA . The fragments were labeled with {gamma-32P}ATP and polynucleotide 5'-hydroxyl-kinase (ATP:5'-dephosphopolynucleotide 5'-phosphotransferase, EC 2.7.1.78) at the 5'-OH groups which were uncovered by alkali treatment . The site is one of three consecutive nucleotides, dA, dA, and dC, located at a unique position . One or a few rA residues were found to be attached to some of the DNA molecules . The transition from the primer RNA to DNA occurs in a region consisting of a segment of five A residues . Both sides of this segment are rich in G and C.

Proc Natl Acad Sci U S A, 1977 May, 74(5), 1851 - 4
Microenvironment of the binding site in the lac carrier protein; Schuldiner S et al.; Studies with a homologous series of (N-dansyl)aminoalkyl-1-thio-beta-D-galactopyranosides containing two to six methylene carbons bridging the galactosyl and dansyl ends of the molecules are described . The compounds were utilized in radioactive and nonradioactive form, and binding of each homologue to membrane vesicles isolated from Escherichia coli ML 308-225 was measured directly by flow dialysis in the presence of D-lactate . The results are compared with the D-lactate-induced fluorescence enhancement observed with each dansylgalactoside and with the ability of N-methylpicolinium perchlorate to quench the fluorescence of the bound homologues . The binding affinity of the lac carrier protein for the probes varies directly with the length of the alkyl linkage, and the same number of binding sites is observed with each homologue . In contrast, however, the increase in fluorescence observed upon binding varies dramatically as the alkyl chain is increased in length, with the fluorescence exhibiting maximal values at two and six methylene carbons and a minimum at four methylene carbons . Furthermore, quenching by N-methylpicolinium perchlorate exhibits an inverse relationship and maximum quenching is observed with the 4 carbon homologue . Possible reasons for this behavior are discussed.

Proc Natl Acad Sci U S A, 1977 May, 74(5), 1811 - 5
Affinity of intact Escherichia coli for hydrophobic membrane probes is a function of the physiological state of the cells; Nieva-Gomez D et al.; The fluorescence parameters of several common membrane probes in the presence of whole E . coli have been examined . The probes included electrically neutral lipophilic molecules N-phenyl-1-naphthylamine, pyrene, and 1,6-diphenyl-1,3,5-hexatriene as well as the negatively charged molecule 8-anilino-1-naphthalene sulfonate . It is demonstrated in each case that certain fluorescence parameters are a function of the state of energization of the cells . All the probes appear to monitor structural changes in the E . coli envelope which accompany the energization and de-energization of the cells . tthe phenomenon is completely reversible as demonstrated by re-energizing anoxic cells by the addition of oxygen, or starved cells by the addition of substrate . All the results are qualitatively consistent with an increased binding of probe by de-energized cells and a subsequent expulsion of probe when the cells are re-energized . A pyrene substituted with a photosensitive group, 1-azidopyrene, has been synthesized . Photolysis in the presence of a suspension of energized E . coli reveals a relatively small amount of probe irreversibly bound to the cells . However, in the presence of cells that have been de-energized the amount of irreversibly bound probe is dramatically increased . This molecule should be useful for localizing the regions of the bacterial envelope that are involved in the structural changes being monitored in these experiments.

J Lipid Res, 1977 May, 18(3), 396 - 9
A sensitive radioenzymatic assay for glycerol and acylglycerols; Schneider PB; A sensitive radioenzymatic assay for glycerol and acylglycerols is described . The assay depends on the quantitative phosphorylation of glycerol to glycerophosphate by glycerol kinase using {gamma-32P}ATP as a substrate . The 32P content of the formed glycerophosphate is determined and gives a measure of the original glycerol content . Acylglycerols can be determined by prior hydrolysis to glycerol . The assay is sensitive to about 0.1 nmol of glycerol and can be extended to 100 nmol . The assay can be applied to the determination of acylglycerols separated by thin-layer chromatography in amounts as low as 0.5 nmol . The assay is particularly useful in the determination of the specific activity of 14C- or 3H-labeled glycerol moeities.

Urology, 1977 May, 9(5), 580 - 5
Clinical and radiographic findings of focally infected polycystic kidneys; Rothermel FJ et al.; Three patients with localized polycystic kidney infections are presented with the pertinent clinical, laboratory, and radiographic findings . Gallium-67 citrate and angiography play an important role in evaluation of these patients . Angiography in particular is valuable in the diagnosis and the exact localization of the inflammatory disease . Localization is extremely important in planning surgical treatment should conservative therapy fail.

J Med Microbiol, 1977 May, 10(2), 225 - 32
A protein factor associated with serum resistance in Escherichia coli; Taylor PW et al.; Immunogel-diffusion studies showed that 60 degree C LiCl extracts of the smooth serum-resistant mutant Escherichia coli strain 17 contained greater amounts of a protein antigen than did extracts of the parent strain LP729 . An extract of strain 17 was fractionated on Sepharose 4B and the protein antigen was found as the only detectable antigen in a number of fractions; sodium dodecyl sulphate-polyacrylamide gel electrophoresis indicated that these fractions contained one major polypeptide band with a molecular weight of 46 000 daltons . We suggest that this protein antigen may be partly responsible for the serum resistance of strain 17 though its presence in other serum-sensitive strains suggests that additional factors are essential for full serum resistance.

J Med Chem, 1977 May, 20(5), 669 - 73
Thymidylate synthetase inhibitors . Synthesis of N-substituted 5-aminomethyl-2'-deoxyuridine 5'-phosphates; Edelman MS et al.; A series of substituted 5-aminomethyl-2'-deoxyuridines was synthesized as analogues of 5-thymidylyltetrahydrofolic acid, a proposed intermediate in the thymidylate synthetase catalyzed reaction . 1-(3,5-Di-O-p-toluoyl-2-deoxy-beta-D-ribofuranosyl)-5-chloromethyluracil (3) was treated with the appropriate amine to give the ester protected 5-aminomethyl nucleoside . Removal of the ester groups was accomplished with anhydrous potassium carbonate in methanol to afford the free beta-nucleoside . In this way 5-(2-dimethylaminoethylaminomethyl)-2'-deoxyuridine (5a), 5-dimethylaminomethyl-2'-deoxyuridine (5b), 5-N-mehtylpiperazinylmethyl-2'-deoxyuridine (5c), and 5-pyrrolidinylmethyl-2'-deoxyuridine (5d) were prepared . Compounds 5a,b,d were converted to the respective 5'-phosphates 6a,b,d . All three compounds were subtrate competitive inhibitors of thymidylate synthetase purified from Escherichia coli, calf thymus, and Ehrlich ascites tumor cells . The most active compound was 6a with KI's of 6,3.1, and 14 micronM observed for the respective enzymes.

J Cell Biol, 1977 May, 73(2), 505 - 19
Site-specific membrane particle arrays in magnesium-depleted Escherichia coli; Weiss RL; The ultrastructure and polypeptide composition of a novel membrane junction in magnesium-starved Escherichia coli are described in this report . Freeze-fracture replicas reveal the junction as a site-specific membrane particle array with four fracture faces . Each junction consists of a cell membrane, a midline zone and a coupled membrane . Membrane particles associated with the junction extend from the hydrophobic region of the cell membrane across the hydrophilic midline zone and into the hydrophobic region of the coupled membrane . After negative staining or after rotary shadowing of freeze-fractured specimens, these particles were seen to consist of two similar but slightly offset bracket-shaped subunits separated by a small space . Optical analysis confirms this structure . Since the apposing membranes are bracketed or linked by their component particles, the name "bracket junction" is proposed for the complex . Methods are described for isolating a membrane fraction enriched in these junctional complexes; the fraction contains a prominent glycoprotein (mol wt 90,000) as well as a number of other components . The bracket junction is compared with the vertebrate gap junction in terms of both structure and possible roles in facilitating the permeation of the cell by small molecules.

J Assoc Off Anal Chem, 1977 May, 60(3), 546 - 62
Methodology for recognition of invasive potential of Escherichia coli; Mehlman IJ et al.; Surveillance for dysentery-related invasive potential in bacteria using the Sereny keratoconjunctivitis test is restricted by expense, time factor, and necessity for confirmation . Primary screening of isolates in a standardized mammalian cell culture system is recommended . Bacteria are grown 20 hr in veal infusion, washed, and resuspended in 20% heat-inactivated fetal bovine serum (FBS) supplemented with 0.12% brain heart infusion and 0.1% bile salts . The HeLa culture is grown 20 hr as a monolayer in chamber slides with 90% minimal essential medium (MEM)-10% FBS . The host culture is infected at a ratio of 10 bacteria/mammalian cell for 3 hr at 35 degrees C . The infection medium is replaced with MEM-FBS supplemented with 300 microng lysozyme and 5 microng gentamycin/ml . The infected monolayer is incubated 5 hr at 35 degrees C to permit intracellular multiplication . Specimens are washed, fixed with methanol, and stained successively with May-Grunwald and Giemsa dyes . Bacteria occur within the cytoplasm if invasion has occurred . The criterion for a positive test is that 1% of the host cells possesses at least 5 bacteria in 2 of 3 trials . Invasiveness is correlated with and possibly preconditioned by cytotoxic principle(s) . Infectivity rates vary from 0 to 30% . The cytopathic effect is noted in 5-50% of HeLa cells . Positive results must be confirmed by the Sereny test.

Eur J Immunol, 1977 May, 7(5), 291 - 7
Genetic selection of mice for quantitative responsiveness of lymphocytes to phytohemagglutinin; Stiffel C et al.; A two-way selection was performed in mice according to the quantitative in vitro response of lymph node lymphocytes to the mitogenic activity of phytohemagglutinin (PHA) . The foundation population was composed of outbred mice produced by reciprocal mating of equal numbers of mice from four different colonies . The selective breeding was carried out by mating of mice at each generation giving the best or the lowest response, respectively . The progressive interline separation produced by 6 generations of selective breeding demonstrates that responsiveness to PHA is submitted to polygenic regulation . The heritability of the character investigated is 0.28 +/- 0.08 . The interline separation is also found with another T mitogen, concanavalin A (Con A) . In spleen cells PHA and Con A produce a similar interline difference . In contrast, the purified protein derivative of tuberculin (PPD) stimulated both lines equally, and E . coli lipopolysaccharide gave only a slightly higher response in high line . This finding implies that our selection based upon response to PHA did not influence B cell function.

Immunology, 1977 May, 32(5), 811 - 7
The humoral immune response of mouse bone marrow lymphocytes in vitro; Ryser JE et al.; Mouse bone marrow (BM) small lymphocytes are shown to contain competent precursors for a primary haemolytic plaque forming cell (PFC) response to heterologous red blood cells and TNP in an in vitro culture system . Their response is dependent on T co-operative factors, which can be provided by irradiated spleen cells activated by concanavalin A or the supernatant of an allogeneic culture, added at the beginning or after 24 h of culture . The frequency of PFC precursors for the response to SRBC is found to be equal or higher in BM than spleen cultures . However, BM lymphocyte cultures stimulated by E . coli lipopolysaccharide show an increase of DNA synthesis but contain only few polyclonal PFC, in contrast to spleen.

Proc Natl Acad Sci U S A, 1977 May, 74(5), 1965 - 8
Role of DNA gyrase in phiX replicative-form replication in vitro; Marians KJ et al.; Preparations containing DNA gyrase activity Gellert, M., Mizuchi, K., O'Dea, M.H . & Nash, H.A . (1976) Proc . Natl . Acad . Sci . USA 73, 3872-3876} have been extensively purified from Escherichia coli . Such fractions, in the presence of ATP and Mg2+, catalyze supertwisting of relaxed circular double-stranded DNA replicative forms of a number of DNAs that results in the formation of superhelical replicative forms . Relaxed phiX174 replicative form (phiX RFIV) is not attacked by the A protein endonuclease coded for by the phiX DNA genome . After exposure to preparations of DNA gyrase, the relaxed phiX174 replicative form is converted to phiX RFI which can then be attacked by the phiX gene A protein and participate in replication of duplex phiX DNA.

J Bacteriol, 1977 May, 130(2), 963 - 4
Dominance of dnaA+ to dnaA in Escherichia coli; Gotfried F et al.; The dominance of dnaA+ to the dnaA508 mutation was complete and was unaffected by the presence of a copy of the chromosomal replication origin on the episome . These results prove that the dnaA gene of Escherichia coli produces a diffusible product.

J Bacteriol, 1977 May, 130(2), 877 - 87
Characterization of lambda Escherichia coli hybrids carrying chemotaxis genes; Silverman M et al.; Molecular cloning techniques were used to construct hybrid Escherichia coli lambda phage and isolate Col E1 factors that carried the cheB region of the E . coli genome . The products of these genes were examined by using a series of deletions in the phage to stimulate specific polypeptide synthesis in ultraviolet-irradiated cells and by using Col factor to program protein synthesis in minicells . Seven flagellar related polypeptides were synthesized . Three of these with apparent molecular weights of 38,000, 28,000, and 8,000 were associated with the cheB region; three polypeptides 63,000, 61,000, and 60,000 were associated with the region that maps between cheB and cheA . These bands were referred to as the triplet group . We suggest that these polypeptides are the same as the methyl-accepting chemotaxis protein described by Kort et al . (Proc . Natl . Acad . Sci . U.S.A . 72:3939-3943, 1975) . Another polypeptide with a molecular weight of 12,000 is associated with the cheA region which also produces at least three gene products . We conclude that the cheA-cheB region in E . coli is complex . Further genetic and biochemical analyses are required to describe all of these products.

J Bacteriol, 1977 May, 130(2), 860 - 8
Analysis of Euglena gracilis chloroplast deoxyribonucleic acid with a restriction endonuclease, EcoRI; Mielenz JR et al.; Cleavage of chloroplast deoxyribonucleic acid (DNA) of Euglena gracilis Z with restriction endonuclease RI from Escherichia coli (EcoRI) yielded 23 bands upon electrophoresis in gels of agarose . Four of the bands contained twice the stoichiometric amount of DNA . One of these bands contained two similarly sized fragments . The sum of the molecular weight of the 24 different fragments equaled the molecular weight of the circular molecule . The restriction fragments had different buoyant densities, with four having distinctly heavy densities in CsCl . Restriction fragments with a high buoyant density were preferentially lost when broken chloroplast DNA was purified by equilibrium density gradient centrifugation . Hybridization of chloroplast ribosomal ribonucleic acid to intact chloroplast DNA determined that there are two cistrons for 16S and 23S ribosomal ribonucleic acid . These two cistrons are located on six restriction fragments, all of which have buoyant densities greater than the intact molecule of chloroplast DNA.

J Bacteriol, 1977 May, 130(2), 846 - 51
Changes in protein synthesis on mitomycin C induction of wild-type and mutant CloDF13 plasmids; Dougan G et al.; Mitomycin C treatment of Escherichia coli K-12 cells containing the nonconjugative plasmid CloDF13 resulted in inhibition of host chromosome protein synthesis and a high rate of synthesis of two CloDF13-specified proteins whose molecular weights correspond to cloacin and immunity protein . Five molecules of immunity protein were synthesized for each cloacin DF13 molecule . Mitomycin C-treated cells containing a copy mutant of CloDF13 made three to four times as much of each protein as cells containing wild-type CloDF13 . CloDF13 plasmids that contained the transposon Tn1 were isolated . Two did not induce after mitomycin C treatment, failing both to inhibit host cell synthesis and to produce the two new proteins . In minicells, they showed reduced CloDF13-specified protein synthesis and produced three Tn1-specified proteins.

J Virol, 1977 May, 22(2), 430 - 45
Specificity of initiation of transcription of simian virus 40 DNA I by Escherichia coli RNA polymerase: identification and localization of five sites for initiation with {gamma-32P}ATP; Lebowitz P et al.; Simian virus 40 (SV40) DNA I was transcribed with Escherichia coli RNA polymerase in the presence of gamma-32P-labeled ribonucleoside triphosphates in order to investigate the specificity of initiation of in vitro transcription . ATP and GTP served as predominant initiating nucleotides, the former being incorporated about twice as much as the latter . Cleavage of {gamma-32P}ATP-labeled SV40 complementary RNA (cRNA) with T1 RNase followed by homochromatographic analysis of the resultant 5' initiation fragments revealed the presence of four specific initiation fragments 6 to 9 nucleotides in length, designated AI, AII, AIIIa, and AIIIb . By means of hybridization of {gamma-32P}ATP-labeled SV40 cRNA to DNA from specific adenovirus 2-SV40 hybrids and specific restriction endonuclease fragments of SV40 DNA before chromatographic analysis, it was possible to identify and determine approximate localizations of five {gamma-32P}ATP initiation sites on the SV40 genome: one in Hin-G close to the Hin-G-B junction, giving rise to the AII fragment, two in the overalpping fragment Hin-A-Hae-A,giving rise to AI and AIII fragments, and two in the fragment Hin-A-Hae-E, also giving rise to AI and AIII fragments . All five sites either fall within or lie near regions of the genome that are cleaved by S1 nuclease and subject to partial alkaline denaturation . These five sites lie on the minus strand of SV40 DNA and initiate RNAs that are copied in a leftward direction . Cleavage of {gamma-32P}GTP-labeled cRNA with pancreatic RNase liberated three major 5' initiation fragments of short length, GI, GII, and GIII, suggesting the presence of three principal GTP initiation sites.

J Bacteriol, 1977 May, 130(2), 676 - 83
Functional capacities and the adenylate energy charge in Escherichia coli under conditions of nutritional stress; Walker-Simmons M et al.; Functional capacities in Escherichia coli cells starved for glucose were examined by comparing protein synthesis, utilization of new substrates, and maintenance of viability with the adenylate energy charge of the culture . When growth ceased because of glucose exhaustion in an E . coli culture, the energy charge dropped from 0.90 to about 0.80 . During this time, the viable-cell count and the capacity for protein synthesis and for induction of new enzymes were maintained only if other substrates were available in the medium . The culture could be maintained for many hours without growth or death if glucose was added slowly; the energy charge in this case stabilized at about 0.80 . A consistent transient decrease in the energy charge to around 0.80, accompanied by a decrease in protein synthesis, was also observed during the adaptation from glucose to other substrates during diauxic growth on glucose and glycerol or lactose.

J Bacteriol, 1977 May, 130(2), 635 - 41
Stringent regulation of the synthesis of a transfer ribonucleic acid biosynthetic enzyme: transfer ribonucleic acid(m5U)methyltransferase from Escherichia coli; Ny T et al.; This paper describes the regulation of a transfer ribonucleic acid (tRNA) biosynthetic enzyme, the tRNA(m5U)methyltransferase (EC 2.1.1.35) . This enzyme catalyzes the formation of 5-methyluridine (m5U, ribothymidine) in all tRNA chains of Escherichia coli . Partial deprivation of charged tRNAVal can be imposed by shifting strains carrying a temperature-sensitive valyl-tRNA ligase from a permissive to a semipermissive temperature . By using two such strains differing only in the allelic state of the relA gene, it was possible to show the tRNA(m5U)methyltransferase to be stringently regulated . Upon partial deprivation of charged tRNAVal, the differential rate of tRNA(m5U)methyltransferase synthesis was found to decrease in a strain with stringent RNA control (relA+), whereas it increased in the strain carrying the relA allele . This increase of accumulation of tRNA(m5U)methyltransferase activity required protein synthesis . Thus, when tRNA is partially uncharged in the cell, the relA gene product influences the expression of tRNA(m5U)methyltransferase gene.

J Bacteriol, 1977 May, 130(2), 620 - 8
Phenethyl alcohol inhibition of sn-glycerol 3-phosphate acylation in Escherichia coli; Nunn WD et al.; In vivo and in vitro experiments were performed to determine how phenethyl alcohol (PEA) inhibits phospholipid synthesis in Escherichia coli . This drug drastically reduced the rate of incorporation of sn-glycerol 3-phosphate into the phospholipids of an sn-glycerol 3-phosphate auxotroph . PEA also reduced the rate of fatty acid incorporation into the phospholipids of a fatty acid auxotroph . The kinetics of PEA inhibition of the rate of incorporation of sn-glycerol 3-phosphate were almost identical to those of PEA inhibition of the rate of fatty acid incorporation into phospholipids . The in vivo experiments suggested that the rate-limiting step(s) in phospholipid biosynthesis inhibited by PEA is at the level of the acylation of sn-glycerol 3-phosphate or beyond this step . PEA inhibited the sn-glycerol 3-phosphate acyltransferase with either palmitoyl coenzyme A or palmitoyl-acyl carrier protein as the acyl donor . This drug, however, had no effect on the cytidine 5'-diphosphate-diglyceride:glycerol 3-phosphate phosphatidyl transferase, cytidine 5'-diphosphate-diglyceride:L-serine phosphatidyl transferase, and acyl coenzyme A:lysophatidic acid acyltransferase . The in vitro findings suggested that PEA inhibits phospholipid synthesis primarily at the level of sn-glycerol 3-phosphate acyltransferase.

Acta Endocrinol (Copenh), 1977 May, 85(1), 55 - 63
Studies on a case of suppurative thyroiditis; Himsworth RL et al.; A patient with suppurative thyroiditis due to infection with Escherichia coli is described . Serial studies of thyroid function were performed . Before the abscess was drained there was no uptake of radio-iodine by the thyroid and there was some spillage of iodinated compounds from the gland . Post-operatively circulating thyroid hormone concentrations declined but were restored at four weeks . Thyroid uptake of radio-iodine was re-established two weeks after operation but there was little secretion of thyroxine at this time . Four weeks after operation the uptake of radio-iodine was greater than normal and thyroxine was being secreted . These changes in thyroid function were not associated with any significant rise in the plasma thyrotrophin concentration.

J Immunol, 1977 May, 118(5), 1852 - 7
Modulation of lipopolysaccharide (LPS)-mediated function by structural differences of two physically distinct fractions of Escherichia coli K235 LPS; Goodman MG et al.; Lipopolysaccharide (LPS), extracted from Escherichia coli K235 by the butanol water technique, was fractionated by gel filtration chromatography into high m.w . (LPS I) and low m.w . (LPS II) fractions . These two forms of LPS were characterized by different densities and chemical compositions . Chemical analysis provided evidence for greater amounts of lipid A and Lipd A-associated protein (LAP) per unit weight associated with LPS II . The biologic activity of the two LPS preparations was compared over a spectrum of different parameters . LPS II was shown to be a more potent mitogen and toxin than LPS I, whereas the two preparations were demonstrated to be of equal activity as polyclonal B cell activators, immunogens, and adjuvants . A modulatory role for the polysaccharide component of the LPS molecule is discussed.

Cancer Res, 1977 May, 37(5), 1438 - 42
Inhibition of the reverse transcriptase of bovine leukemia virus by antibody in sera from leukemic cattle and immunological characterization of the enzyme; Wuu KD et al.; Sera from some leukemic cattle contain an antibody that inhibits the reverse transcriptase activity of the bovine leukemia virus . The antibody is not directed against the synthetic template or the major internal and envelope viral antigens . The antibody failed to inhibit the DNA polymerases of the murine leukemia virus, simian sarcoma-associated virus, avian myeloblastosis virus, or Escherichia coli . Conversely, the bovine leukemia virus enzyme was not inhibited by antibody against the reverse transcriptases of other C-type viruses . These findings agree with previous results showing that the major internal bovine leukemia virus protein lacks the known interspecies- and intraspecies-specific antigenic determinants indentified in the homologous proteins of other oncornaviruses.

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