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