Biochim Biophys Acta, 1975 Oct 15, 407(3), 320 - 4 Characterization of the particles produced by exposure of ribosomal subunits to urea; Langer JA et al.; When Escherichia coli 50-S ribosomal subunits are treated with increasing concentrations of urea partial deproteination occurs . Furthermore, we observed that the number of sulfhydryl groups which react with Ellman's reagent is a sigmoidal function of the urea concentration . These results are similar to those previously reported for the 30-S subunit (Acharya, A.S . and Moore, P.B . (1973) J . Mol . Biol . 76, 207-221) . For both subunits we identify the proteins which dissociate (split proteins) or are recoverable in a ribonucleoprotein particle (core proteins) under the action of 6 M urea in a buffer of moderate ionic strength. Biochim Biophys Acta, 1975 Oct 15, 407(3), 273 - 82 Progress in the resolution of the cytoplasmic membrane DNA initiation complex of Escherichia coli; Gomez-Eichelmann MC et al.; The attachment of the bacterial chromosome to the cytoplasmic membrane in Escherichia coli was studied . The initiator DNA was specifically labeled and the outer and cytoplasmic membranes were separated in a step sucrose gradient . The labeled DNA was localized mainly in the cytoplasmic membrane fraction . The DNA . cytoplasmic membrane complex was isolated from cells uniformly labeled with {Me-3H}thymidine, solubilized with deoxycholate and chromatographed on Sepharose 4B . A high percent of the labeled DNA was excluded in the void volume but a small fraction eluted associated with the second protein elution peak . The isolation of such a DNA . cytoplasmic membrane protein complex, suggests useful strategies for future studies about the molecular components of the initiation complex in E . coli. Eur J Biochem, 1975 Oct 15, 58(2), 611 - 9 Envelope-bound N-acetylmuramyl-L-alanine amidase of Escherichia coli K 12 . Purification and properties of the enzyme; van Heijenoort J et al.; N-Acetylmuramyl-L-alanine amidase activity was detected in Escherichia coli K 12 by usine N-acetylmuramyl-L-alanyl-gamma-D-glutamyl-(L)-meso-{3H}diaminopimelic acid as a radioactive substrate . This activity cleaves the amide bond between the residues of N-acetylmuramyl acid and L-alanine . It was readily obtained in a soluble form either by mechanical disruption of the cells or by spheroplast formation . In the latter case the release of most of the activity into the sucrose medium seems to indicate that it is either periplasmic or associated with the outer membrane of the envelope of E . coli K 12 . The enzyme was purified to near homogeneity . A molecular weight of 39 000 was determined by gel filtration and confirmed by polyacrylamide gel electrophoresis . Further characterisation of this N-acetylmuramyl-L-alanine amidase activity was carried out by investigating several of its properties. Eur J Biochem, 1975 Oct 15, 58(2), 501 - 10 The importance of Escherichia coli ribosomal proteins L1, L11 and L16 for the association of ribosomal subunits and the formation of the 70-S initiation complex; Kazemie M; 50-S subunits were washed with LiCl solutions of different concentrations . After washing with 1 M LiCl solution the particles lost their ability to attach either to 30-S subunits or to the AUG - 30-S subunit - fMet-tRNAfMet complex or to a poly(U) - 30-S subunit - Phe-tRNAPhe complex . Those proteins which were removed by LiCl were fractionated on a Sephadex G-100 column . Of the fractionated proteins only the combinations L1 and L11 or L1 and L16 were essential for the association of 50-S 1.0 cores (particles prepared by washing 50-S subunits in 1.0 M LiCl) with 30-S subunits . These three proteins were also required for the formation of a stable complex between 50-S 1.0 cores, mRNA, 30-S subunits and aminoacyl-tRNA. Eur J Biochem, 1975 Oct 15, 58(2), 383 - 95 The use of primed synthesis by DNA polymerase I to study an intercistronic sequence of phiX-174 DNA; Donelson JE et al.; A decadeoxynucleotide complementary to ten nucleotides in the major ribosome-protected fragment of phiX-174 plus-strand DNA has been chemically synthesized and used as a primer for DNA polymerase I on phiX-174 plus-strand DNA as template . The sequence of the first 40 nucleotides incorporated onto the decadeoxynucleotide has been determined . This sequence extends further the sequence of the intercistronic region preceding gene G and shows the presence of another termination codon . The sequence was determined by using manganese as the activating cation for DNA polymerase I which allows ribonucleotides to be incorporated as well as deoxyribonucleotides . The ribo-substituted product was then cleaved specifically at the ribonucleotide residues to generate a series of overlapping ribo-terminated fragments whose sequences were sufficient to determine the complete sequence of the first 40 nucleotides . No evidence for misincorporation by DNA polymerase I in the presence of manganese was detected. Eur J Biochem, 1975 Oct 15, 58(2), 359 - 66 Phage Qbeta replicase: cell-free synthesis of the phage-specific subunit and its assembly with host subunits to form active enzyme; Happe M et al.; Cell-free translation of Qbeta RNA and subsequent partial purification of the enzyme resulted in replicase activity . From 0.5 to 1.5% of all R chains synthesised were found in the 7-S replicase complex . The presence in the 7-S complex of the host subunits of authentic replicase, i (= S1) and EF-Ts, was shown by the effect of antisera directed against ribosomal protein S1 and EF-Ts, respectively . Furthermore, the presence of EF-Ts was demonstrated by thermal denaturation of in vitro replicase made by a cell extract from an Escherichia coli mutant with a thermolabile EF-Ts . In vitro replicase did not assemble spontaneously during protein synthesis but was formed upon subsequent purification . Assembly could be induced by ammonium sulphate precipitation (60% saturation) alone . It is concluded that the functional phage-coded subunit synthesised in vitro recognises i and the EF-Tu - EF-Ts complex among a mixture of host proteins. Eur J Biochem, 1975 Oct 15, 58(2), 269 - 72 On the dissociation and association of Escherichia coli ribosomes; Walters H et al.; The dissociation and association behaviour of 70-S ribosomes of Escherichia coli has been studied . It has been shown that the dissociation-association reaction can be both a real dynamic equilibrium and a non-equilibrium reaction, dependent upon the ionic conditions of the solvent . At relatively high ionic strength (I = 0.15 M or more) the dissociation-association reaction is an equilibrium reaction, whereas at lower ionic strength (I = 0.1 M or less) there is no dynamic equilibrium between 70-S ribosomes and its subunits . In the latter case a hysteresis in the dissociation-association reaction is observed . Whether there is a dynamic equilibrium or not can be demonstrated by a single centrifugation experiment, using the analytical ultracentrifuge. Eur J Biochem, 1975 Oct 15, 58(2), 303 - 13 Replication of colicinogenic factor E1 DNA in plasmolysed Escherichia coli cells . Coupling of DNA replication and RNA synthesis; Staudenbauer WL; Plasmolysed chloramphenicol-treated Escherichia coli cells carrying the colicinogenic factor E1 utilize deoxynucleoside triphosphates for the semi-conservative synthesis of Col E1 DNA . Col E1 DNA replication in plasmolysed cells can be dissociated into two temporally separated processes: (a) a rifampicin-sensitive RNA synthesis, which is stimulated by adenosine 3':5'-monophosphate (cyclic AMP) and requires all four ribonucleoside triphosphates and (b) an ATP-dependent DNA synthesis, which is inhibited by arabinosylnucleoside triphosphates and sulfhydryl-blocking reagents . Thes two processes exhibit different sensitivities to inhibition by polyamines and actinomycin D. Biochim Biophys Acta, 1975 Oct 15, 407(3), 357 - 64 Inhibition of leucyl-tRNA synthetase in Escherichia coli by the cytostatic 5,8-dioxo-6-amino-7-chloroquinoline; Ogilvie A et al.; At concentrations of 1-1.6 mug/ml, 5,8-dioxo-6-amino-7-chloroquinoline causes auxotrophy for leucine in Escherichia coli MRE 600 . With increasing concentrations of this quinone additional amino acids are required for growth . The amount of leucine in the pool of free amino acids is not decreased after treatment of E . coli with the quinone . Transfer RNALeu, however, is charged with leucine less than 10% in quinone-treated cells of E . coli, whereas in control cells the degree of aminoacylation is about 85% . From these data we conclude that the quinone causes auxotrophy for leucine by interacting with the charging process of tRNALeu . Quinone was found to inhibit leucyl-tRNA synthetase activity in purified extracts of E . coli with E . coli tRNA as substrate. J Biol Chem, 1975 Oct 10, 250(19), 7759 - 65 Inactivation of normal beta-D-galactosidase by antibodies to defective forms of the enzyme; Roth RA et al.; A counterpart of the antibody-mediated activation of genetically defective enzymes is reported here . Antibodies elicited by certain mutant forms of beta-D-galactosidase (EC 3.2.1.23) of Escherichia coli were found to inactivate the normal form of the enzyme . (Antibodies elicited by normal beta-D-galactosidase do not affect the enzyme's catalytic activity.) We present evidence that the inactivating antibodies are directed against one or a few determinants of the enzyme . The level of inactivation caused by the antibodies was independent of temperature below 25 degrees and increased with temperature above 25 degrees . The inactivation was proportional to the concentration of antiserum until a maximum level of 50% inactivation was reached . Antibodies capable of inactivating up to 87% of the activity were obtained after the antiserum was partially absorbed in an affinity column . This antibody preparation showed a 10-fold enrichment of inactivating antibodies over other antibodies direct against the enzyme . The antibody-mediated inactivation caused a reduction in the Vmax of beta-D-galactosidase without affecting the apparent Km of the enzyme . In contrast to antibodies to normal beta-D-galactosidase, inactivating antibodies changed the response of the enzyme to cations . To explain these results, we present a model in which there is a temperature-dependent equilibrium between two active forms of beta-D-galactosidase . Inactivation results from a conformational change induced by the binding of inactivating antibodies to only one of these two forms. J Biol Chem, 1975 Oct 10, 250(19), 7771 - 9 Protease II from Escherichia coli . Purification and characterization; Pacaud M et al.; We have previously demonstrated the existence of two types of endopeptidase in Escherichia coli . A purification procedure is described for one of these, designated protease II . It has been purified about 13,500-fold with a recovery of 24% . The isolated enzyme appears homogeneous by electrophoresis and gel filtration . Its molecular weight is estimated by three different methods to be about 58,000 . Its optimal pH is around 8 . Protease II activity is unaffected by chelating agents and sulfhydryl reagents . Amidase and proteolytic activities are stimulated by calcium ion, which decreases the enzyme stability . Like pancreatic trypsin, this endopeptidase catalyses the hydrolysis of alpha-amino-substituted lysine and arginine esters . It appears distinct from the previously isolated protease I, which is a chymotrypsin-like enzyme . The apparent Michaelis constant for hydrolysis of N-benzoyl-L-arginine ethyl ester is 4.7 X 10(-4) M . The esterase activity is inhibited by diisopryopylphosphorofluoridate (Ki(app) equals 2.7 X 10(-3) M) and tosyl lysine chloromethyl ketone (Ki(app) equals 1.8 X 10(-5) M), indicating that serine and histidine residues may be present in the active site . However, protease II is insensitive to phenylmethanesulfonyl fluoride and several natural trypsin inhibitors . Its amidase and esterase activities are competitively inhibited by free arginine and aromatic amidines . The proteolytic activity measured on axocasein is very low . In contrast to trypsin, protease II is without effect on native beta-galactosidase . It easily degrades aspartokinase I and III . Nevertheless both enzymes are resistant to proteolysis in the presence of their respective allosteric effectors . These results provide further evidence that such differences in protease susceptibility can be related to the conformational state of the substrate . The possible implication of structural changes in the mechanism of preferential proteolysis in vivo, is discussed. J Biol Chem, 1975 Oct 10, 250(19), 7687 - 92 Characterization of an active transport system for calcium in inverted membrane vesicles of Escherichia coli; Tsuchiya T et al.; The energy-dependent uptake of calcium by inverted membrane vesicles of Escherichia coli was investigated . Methods for preparation and storage of the vesicles were devised to allow for the maximal activity and stability of the calcium transport system . The pH and temperature optima for the reaction were observed to occur at pH 8.0 AND 30 DEGREES, RESPECTIVELY . The eft was found that the extent of the reaction depended on the presence of phosphate or oxalate . Phosphate was found to enter the vesicles at a rate slower than that of calcium . A Ca2+:Pi ratio of approximately 1.5 was found, suggesting formation of Ca3(PO4)2 . Monovalent cations stimulated calcium uptake, with the order of effectiveness being K+ is greater than Na+ is greater than Li+ is greater than NH4+ . Inhibition was found with certain divalent cations, but these also inhibited the electron transport chain . Of the divalent cations examined only Mg2+ and Sr2+ inhibited calcium transport without a corresponding inhibition of respiration . Calcium transport exhibited biphasic Kinetics, with a low affinity system and a high affinity system . The low affinity system showed a Km of 0.34 mM and a Vmax of 85 nmol/min/mg of protein . The kinetic constants of the high affinity system were 4.5 muM and 2 nmol/min/mg of protein . The energy for calcium transport could be derived from the electron transport chain by oxidation of NADH, D-lactate, and succinate, in order of their effectiveness . Respiration-driven calcium transport was inhibited by inhibitors of the electron transport chain and by uncouplers of oxidative phosphorylation . ATP could also be used to supply enerty for calcium transport . The ATP-driven reaction was inhibited by inhibitors of the Mg2+ATPase and by an antiserum prepared against that protein, demonstrating that that enzyme is involved in the utilization of ATP for active transport in inverted vesicles. Biochim Biophys Acta, 1975 Oct 10, 408(1), 47 - 57 Isolation and properties of Escherichia coli ATPase mutants with altered divalent metal specificity for ATP hydrolysis; Thipayathasana P; A method was devised for isolation of large numbers of energy-transducing ATPase (coupling factor) mutants based on a modification of the procedure of Hong and Ames (Hong, J . and Ames, B . N . (1971) Proc . Natl . Acad . Sci . U.S . 68, 3158-3162) for localized mutagenesis of any small region of the bacterial chromosome using transducing phages . The principle of this procedure is to mutate P1-transducing phage particles carrying the ATPase genes (Unc (uncoupled) DNA) using the strong chemical mutagen hydroxylamine . By transducing ilv- auxotrophs, a marker closely linked to Unc, to prototrophs, mutated Unc DNA can be introduced into the chromosome . We have used this method in conjunction with suitable selection procedures to isolate about 90 Unc- strains which have been classified by physiological, genetic, and biochemical criteria into three different phenotypes (Unc A, B, D) . Mutants of the Unc D phenotype which were studied in detail were found to have the following properties: (1) aerobic growth yields on glucose are considerably lower than the wild type; growth occurs on glucose under anaerobic conditions; (2) Unc D lesions map near the ilv operon; (3) O2 uptake is comparable to the rate of wild type; (4) vesicles catalyze respiratory-dependent transhydrogenation, but show very low levels of Ca2+ ATP-dependent transhydrogenation; Mg2+ is ineffective; (5) oxidative phosphorylation is almost completely blocked irrespective of which metal ion is used; (6) the specific activity of ATPase is only about 20% of the wild type: (7) purified ATPase was found to have a marked specificity for Ca2+ as a divalent metal for ATP hydrolysis . A summary of properties of the new Unc mutants is discussed. J Biol Chem, 1975 Oct 10, 250(19), 7668 - 74 Active site stoichiometry of L-phenylalanine: tRNA ligase from Escherichia coli K(-10); Bartmann P et al.; The existence of two active siter per molecule of L-phenylalanine:tRNA ligase from Escherichia coli K(-10) has been demonstrated by isolation of the E-aminoacyl adenylate and tel filtration and the nitrocellulose filter assay at pH 5.0 revealed the same stoichiometry for the E-tRNAPhe comples as protection against degradation by snake venom phosphodiesterase and equilibrium gel filtration at pH 7.5 . Using a fluorescence titration technique, it was found that the dissociation constant for ligase-tRNAPhe complex is decreased 20-fold when the hydrogen ion concentration is changed from pH 6.0 to pH 5.0 . The existence of two active sites binding the aminoacyl adenylate intermediate was demonstrated by gel filtration and retention on DEAE-cellulose filters . "Burst" experiments indicated that two sites were involved in a rapid ATP consumption at conditions of catalytic amino acid activation . Furthermore, it was observed that the activated amino acid could be transferred from both sites to cognate tRNA. Biochim Biophys Acta, 1975 Oct 9, 404(2), 180 - 7 Effect of L-azetidine 2-carboxylic acid on growth and proline metabolism in Escherichia coli; Grant MM et al.; The effects of L-azetidine 2-carboxylic acid on growth and proline metabolism in a proline-requiring auxotroph of Escherichia coli are described . The homologue inhibited growth of the wild type and it, alone, did not substitute effectively for proline as a growth supplement for the mutant . In medium containing 0.05 mM proline, the addition of increasing amounts of homologue progressively inhibited growth of the wild type but stimulated growth of the mutant at homologue: proline ratios of 10 : 1 and 50 : 1 . This suggested that the homologue exerted a "sparing effect" on proline in the mutant . The incorporation of L-{U-14C}proline and L-{3H}azetidine 2-carboxylic acid into hot trichloroacetic acid-insoluble material in the mutant was measured . Amino acid analysis of the insoluble material from cells incubated with radiolabeled proline alone revealed that proline was partially degraded and metabolized to other amino acids prior to incorporation into protein . The addition of unlabeled homologue to the incubation medium significantly reduced proline catabolism, suggesting that the homologue exerted a sparing effect on proline in this mutant . In medium containing unlabeled proline and radiolabeled L-azetidine 2-carboxylic acid, the homologue was incorporated both intact and partially degraded prior to incorporation into protein . Alanine was the major L-azetidine 2-carboxylic acid catabolite. Biochim Biophys Acta, 1975 Oct 9, 404(2), 333 - 40 Phosphonopyruvic acid: A probable precursor of phosphonic acids in cell-free preparation of Tetrahymena; Horiguchi M et al.; 1 . In cell-free preparations of Tetrahymena, doubly labelled {32P}phosphoenol-{3-14C}pyruvate gives rise to 2-aminoethylphosphonate and 2-amino-3-phosphonopropionate, labelled with the two isotopes in the same ratio as the starting compound . The result is consistent with an intra-molecular rearrangement of phosphoenolpyruvate in the biosynthetic sequence of carbon-phosphorus bond formation . 2 . Incubation of {32P}phosphoenolpyruvate with the same preparation, followed by treatment with 2,4-dinitrophenylhydrazine, yielded labelled hydrazones . When these were subjected to hydrogenolysis, the radioactivity was recovered in 2-aminoethylphosphonate and 2-amino-3-phosphonopropionate, suggesting that 2-phosphonoacetaldehyde and 3-phosphonopyruvic acid were probable precursors of the aminoalkylphosphonic acids . 3 . Radioactivity from 2-amino-3-phosphono-{3-14C}propionic acid was incorporated into 2-aminoethylphosphonic acid, but incorporation of the radioactivity into lipids was negligible. Biochemistry, 1975 Oct 7, 14(20), 4552 - 8 DNA-binding chromosomal non-histone proteins . Isolation, characterization, and tissue specifity; Chiu JF et al.; A fractionation schedule is described which allows the isolation of a group of chromosomal non-histone proteins (NP) with affinity for DNA . In polyacrylamide gel electrophoresis these proteins isolated from rat liver are represented principally by a group of low molecular weight polypeptides . The NP fraction comprises about 2-4% of the total chromatin protein content in rat liver or Novikoff hepatoma . Experiments in vivo and in vitro revealed that the NP proteins do not incorporate significant amounts of 32P . Complexes of the chromosomal proteins NP with homologous DNA are immunologically tissue specific and the specificity can be transferred by reconstituting the NP proteins from one tissue to the residual chromatin from another. Biochemistry, 1975 Oct 7, 14(20), 4482 - 6 Cobalt(III) labeled aspartokinase-homoserine dehydrogenase of Escherichia coli; Ryzewski C et al.; The kinase activity of the threonine-sensitive aspartokinase-homoserine dehydrogenase enzyme complex of Escherichia coli was selectively inactivated by Co(III) incorporation . Incubation of the enzyme with Co(II) in the presence of oxygen or H2O2 resulted in incorporation of one Co(III) per subunit . The cobalt(III) bound to the enzyme was not removable by dialysis and presumably results from formation of "inert" coordination complexes with ligands contributed by the enzyme . Cobalt was released from the enzyme by incubation with dithiothreitol but not by metal chelating agents . The Co(III)-labeled enzyme was aspartokinase inactive but still retained 60% of its original homoserine dehydrogenase activity . Studies of the time course of inactivation showed aspartokinase inactivation paralleled Co(III) incorporation . The residual dehydrogenase activity of aspartokinase inactive enzyme was still inhibited by threonine Thus, Co(III) incorporation seems to result in a specific inactivation of kinase activity which permits enumeration of the number of aspartokinase sites . Limited alpha-chymotrypsin digestion of Co(III)-enzyme produced homoserine dehydrogenase-active fragments devoid of Co(III), further confirming the specificity of the labeling procedure . Aspartokinase inactivation obtained without concomitant desensitization of homoserine dehydrogenase to threonine inhibition suggests that kinase active site integrity is not required for threonine binding and inhibition of homoserine dehydrogenase. Biochemistry, 1975 Oct 7, 14(20), 4414 - 20 Escherichia coli stringent factor binds to ribosomes at a site different from that of elongation factor Tu or G; Richter D et al.; The binding of Escherichia coli stringent factor to ribosomes has been studied; the reaction depends on 50S and 30S ribosomal subunits and poly(U) as messenger RNA . The ribosome-stringent factor complex is formed at 5-10 mM magnesium acetate; NH4 ions are inhibitory . Binding of the stringent factor to the 70S-mRNA complex is not stimulated by uncharged tRNA . The ribosomal binding site(s) for the stringent factor does not overlap with the one known for the elongation factor Tu (EF-Tu) or G (EF-G) . Ribosomes carrying either EF-Tu or EF-G are active in binding the stringent factor; however, they are inactive in synthesizing guanosine 5'-triphosphate 3'-diphosphate (pppGpp) and guanosine 5'-diphosphate 3'-diphosphate (ppGpp) . The latter result is due to the blockage of the ribosomal acceptor site by the aminoacyl-tRNA and/or elongation factors . That stringent and elongation factors do not compete for identical ribosomal region(s) is supported by: (1) the reverse experiments where ribosomes charged with the stringent factor are fully active in EF-Tu or EF-G dependent functions; (2) ribosomes that lack the two ribosomal proteins L7 and L12 known to be essential for EF-Tu and EF-G functions bind the stringent factor and are active in synthesizing pppGpp and ppGpp. Mol Gen Genet, 1975 Oct 3, 140(3), 253 - 74 The stoichiometry of the ribosomal proteins of Escherichia coli; Hardy SJ; A ribosome preparation from E . coli made without stringent washing procedures has been shown to contain the same relative amounts of nearly all the ribosomal proteins as ribosomes in intact cells . Stoichiometric measurements on all the proteins of this preparation except for L8, L20, L31 and L34 have been made using an isotope dilution technique . When the scatter of the values obtained, the uncertainty in the molecular weights, and the losses occurring during extraction are taken into account, none of the proteins except L7/L12 is present at a level significantly different from one molecule per ribosome . There are multiple copies of L7/L12 . These data suggest that the ribosomes of Escherichia coli are homogeneous in vivo. Mol Gen Genet, 1975 Oct 3, 140(3), 221 - 30 Mutagenic DNA repair in Escherichia coli . II . Factors affecting loss of photoreversibility of UV induced mutations; Doubleday OP et al.; The photoreversibility of UV-induced mutations to Trp+ in strain Escherichia coli WP2 uvrA trp (unable to excise pyrimidine dimers) was lost at different rates during incubation in different media . In Casamino acids medium after a short initial lag, photoreversibility was lost over about one generation time; in minimal medium with tryptophan, photoreversibility persisted for more than two generations; in Casamino acids medium with pantoyl lactone photoreversibility was lost extremely slowly . The rate of loss of photoreversibility was unaffected by UV dose in either Casamino acids medium or in minimal medium . The same eventual number of induced mutants was obtained when cells were incubated for two generations in any of the three media before being transferred to selective plates supplemented with Casamino acids . Thus in each the proportion of cells capable of giving rise to a mutant was the same and only the rate at which these cells did so during post-irradiation growth varied, suggesting that there might be a specific fraction of pyrimidine dimers at a given site capable of initiating a mutagenic repair event, and that the size of this fraction is dose dependent . Segregation experiments have shown that error-prone repair appears to occur once only and is not repeated in subsequent replication cycles, in contrast to (presumed error-free) recombination repair . The results are discussed in the light of current models of UV mutagenesis. Bull Los Angeles Neurol Soc, 1975 Oct, 40(4), 140 - 4 Ventriculogastrostomy, an alternative means for CSF diversion: a preliminary study; Weiss MH et al.; The feasibility of gastric CSF diversion in the management of hydrocephalus is evaluated in laboratory and clinical settings . A technique for ventriculogastrostomy is described and evaluated initially in 8 mongrel dogs . All distal shunts remained patent to the time of sacrifice . None of the animals exhibited leakage of gastric contents around the tubing . Cultures of the components of the shunting system and gastric mucosa were sterile . Clinical evaluation in a 3 week-old child is discussed . The trial was terminated at 3 weeks postsurgery because of the occurrance of an E . coli ventriculitis which was considered to be secondary to a preoperative conjunctivitis in which the organism was identical . It is concluded that there is good experimental evidence to support the concept of effectiveness of ventriculogastrostomy in the treatment of selected cases of hydrocephalus. Z Gastroenterol, 1975 Oct, 13(6), 573 - 6 {Studies on the cytotoxicity of lymphocytes in ulcerative colitis}; Kovacs A et al.; Cytotoxic action was examined with 51 Cr labelled fetal human colon epithelial cell cultures -- using lymphocytes of healthy test persons and of patients suffering from ulcerous colitis and colon carcinoma . There was no significant difference as to the cytotoxic action of the lymphocytes between the three above-mentioned groups; these results are in contrast to those obtained by previous research . This fact is considered to be due to the different methods applied . In the cases with ulcerous colitis E . coli or E . coli with CEA together formed the antigen against which complex immuno-processes are generated. Nucleic Acids Res, 1975 Oct, 2(10), 1931 - 9 Distribution of the modified nucleoside Q and its derivatives in animal and plant transfer RNA's; Kasai H et al.; The modified nucleoside, 7-(4,5-cis-dihydroxy-1-cyclopenten-3-yl-aminomethyl)-7-deazaguanosine, designated as Q, and its derivative, Q*, were found in tRNA's from various organisms, including several mammalian tissues, other animals such as starfish, lingula and hagfish, and wheat germ . Q isolated from rat liver tRNA was found to be identical with E . coli Q by mass spectrometry and thin-layer chromatography . Thus the rare modified nucleoside Q originally isolated from E . coli tRNA, is widely distributed in various organisms . Analysis of the mass spectrum of Q* suggested that it has a different side chain from Q. Biochim Biophys Acta, 1975 Oct 1, 407(2), 125 - 32 Uterine chromatin template activity during the early stages of pregnancy in the rat; O'Grady JE et al.; 1 . The template activity of chromatin prepared from rat uterine nuclei during dioestrus, oestrus and the first 7 days of pregnancy has been examined . 2 . The DNA, RNA, histone and non-histone protein contents of uterine chromatin remained constant during early pregnancy . 3 . The rate of RNA synthesis on Day 1 uterine chromatin was 8.61 +/- 0.59 (mean +/- S.E.) pmol of UMP incorporated/mg DNA per 10 min . When compared with DNA prepared from rat liver nuclei, 13.20 +/- 0.27% (mean +/- S.E.) of the Day 1 chromatin DNA was available for transcription by Escherichia coli RNA polymerase . 4 . Uterine chromatin from rats in early dioestrus had significantly less template activity than during oestrus . 5 . Chromatin prepared from whole uterus on Day 5 and from implantation sites on Days 6 and 7 of pregnancy had a significantly higher template activity than chromatin obtained from uteri on Day 1 . Chromatin from interimplantation tissue on Day 6 had a lower template activity than that from uteri on Day 1 . 6 . RNA - DNA hybridisation of RNA transcribed from chromatin obtained on Days 2, 5 and 7 of pregnancy showed that RNA transcribed from Day 5 chromatin obtained species not present (or present in very small amounts) in RNA transcribed by chromatin from uteri on Day 2 and from implantation tissue on Day 7 of pregnancy . 7 . The results are discussed in relation to the cellular changes occurring in the stroma immediately before implantation and it is postulated that the appearance of a new species of RNA on Day 5 is related to the preparation of the stromal cells for decidualisation. Cancer Res, 1975 Oct, 35(10), 2830 - 5 Ethionine-induced changes in rat liver transfer RNA methylation; Wainfan E et al.; We have confirmed the finding by Rajalakshmi that transfer RNA (tRNA) from livers of ethionine-treated rats can act as a substrate for homologous tRNA-methylating enzymes in vitro . This methyl-deficient tRNA from liver can be methylated in vitro by enzymes from normal or ethionine-treated rats . The in vitro inhibition of tRNA methylation that follows ethionine treatment can be at least partially relieved in vitro . The liver extracts from ethionine-treated animals contained a low-molecular-weight inhibitor of tRNA methylation . Dialysis of enzyme preparations from ethionine-treated, but not control, rats resulted in large increases in tRNA methylase activity, with either Escherichia coli or homologous tRNA's as substrate . Furthermore, the tRNA methylase activity of control rat liver enzyme extracts was greatly depressed by dialysate from liver homogenates of ethionine-treated rats . After 5 days of ethionine administration the liver tRNA methylase activities were significantly higher than those of control preparations despite the continued presence of the dialyzable inhibitor(s) . The liver tRNA's from these animals were poorer methyl acceptors than those from 3-day-treated rats, although still better than tRNA's from untreated rats . These observations have been interpreted to indicate that ethionine causes the accumulation in the liver of inhibitors of tRNA methylation . Early in the course of ethionine administration, methyl-deficient tRNA can be isolated . When the period of ethionine treatment is extended, the organism attempts to maintain homeostasis by production of increased amounts of tRNA-methylating enzymes . The increased quantities of these enzymes are able to overcome, at least partially, the effects of the inhibitors and to decrease the extent to which methyl-deficient tRNA is produced. Ann Microbiol (Paris), 1975 Oct-Nov, 126(3), 273 - 85 Sythesis and turnover of phospholipids in Escherichia coli cells with inhibited DNA synthesis; Michel G et al.; The relative amounts of individual phospholipids are not greatly affected in E . coli B cells inhibited with mitomycin C, nalidixic acid or hydroxyurea . Turnover and synthesis of phospholipids continue in the first 40 minute period of treatment with nalidixic acid and then cease . Arrest of phospholipid synthesis is not coupled with protein synthesis since beta-galactosidase is still induced after cessation of phospholipid metabolism . The cells, washed free of drug, resume growth as well as synthesis and turnover of phospholipids . During the lag phase preceding the renewal of cell division, a decrease in phosphatidylglycerol content and an increase in cardiolipin content are observed . The ratio of these phospholipids is then comparable with that found in normal non-dividing cells . We suggest that the variation in the relative contents of cardiolipin and phosphatidylglycerol might be related to the cell division process. Lab Anim, 1975 Oct, 9(4), 329 - 36 The use of the pig as an animal model to study problems associated with low birthweight; Cooper JE; Attention is drawn to some of the consequences associated with the postnatal development of low birthweight human infants and the use of the piglet as a model to study such problems . A description is given of the production and methods of rearing of these animals along with an outline of studies currently in progress. J Reprod Fertil Suppl, 1975 Oct, (23), 605 - 10 Infection of the horse fetus; Platt H; Many infections of the equine placenta and fetus result from ascending spread along the cervical canal . Most abortions due to infection occur during the later stages of pregnancy and the possible effects of intrauterine infection on the developing fetus and young foal are discussed. Biochem Genet, 1975 Oct, 13(9-10), 635 - 41 The molecular mechanism of benzimidazole mutagenicity: in vitro studies on transcription and translation; Seiler JP; Benzimidazoles are weak mutagens acting through base substitutions; they are incorporated into nucleic acids . Experiments with deoxyribohomopolymers as templates demonstrated that benzimidazole nucleoside triphosphate is polymerized by RNA polymerase only in the presence of poly dC, i.e., instead of guanine . In plasmolyzed Escherichia coli cells, benzimidazole ribonucleoside diphosphate is polymerized by polynucleotide phosphorylase and can, after blocking of the normal mRNA synthesis with actinomycin D, be used as a messenger for polypeptide formation . The addition of radioactive amino acids to this system showed that benzimidazole is not read preferentially as guanine, as would have been expected from the RNA polymerase results . Instead, the reading was position dependent and brnzimidazole is recognized (1) in the first codon position as adenine, (2) in the second as purine, and (3) in the third possibly only as base . Benzimidazole mutagenicity is thus explained as a G in equilibrium A transition. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3961 - 5 Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene; Grunstein M et al.; A method has been developed whereby a very large number of colonies of Escherichia coli carrying different hybrid plasmids can be rapidly screened to determine which hybrid plasmids contain a specified DNA sequence or genes . The colonies to be screened are formed on nitrocellulose filters, and, after a reference set of these colonies has been prepared by replica plating, are lysed and their DNA is denatured and fixed to the filter in situ . The resulting DNA-prints of the colonies are then hybridized to a radioactive RNA that defines the sequence or gene of interest, and the result of this hybridization is assayed by autoradiography . Colonies whose DNA-prints exhibit hybridization can then be picked from the reference plate . We have used this method to isolate clones of ColE1 hybrid plasmids that contain Drosophila melanogaster genes for 18 and 28S rRNAs . In principle, the method can be used to isolate any gene whose base sequence is represented in an available RNA. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3947 - 51 Polypeptide chain initiation in eukaryotes: initiation factor MP in Artemia salina embryos; Filipowicz W et al.; The activity of IF-MP, a polypeptide chain initiation factor that forms a ternary complex with eukaryotic initiator Met-tRNA and GTP and promotes binding of the initiator to 40S ribosomes, is very low in undeveloped Artemia salina embryos but increases over 20-fold following resumption of development upon hydration of the cysts . The factor is present in both the ribosomal salt wash and high-speed-supernatant . Its specific activity is 50 times higher in the wash but its total activity is only about twice as high in the wash as in the supernatant . As is true of IF-MP from other eukaryotic sources, the A . salina factor is specific for eukaryotic Met-tRNAi and sensitive to SH-reagents, and its activity is GTP dependent. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3921 - 4 Role of leucyl-tRNA synthetase in regulation of branched-chain amino-acid transport; Quay SC et al.; The regulation of the transport of leucine, isoleucine, and valine in Escherichia coli B/r was studied in a mutant with a complete deletion of the leucine biosynthetic operon and a temperature-sensitive leucyl-tRNA synthetase {L-leucine:tRNALeu ligase (AMP-forming), EC 6.1.1.4} . Under conditions of excess leucine and a functional leucyl-tRNA synthetase transport activity was repressed . Shifting the culture to a temperature at which the activation of leucine to an appropriate tRNA species became growth-rate-limiting led to a large increase in the high-affinity transport of leucine, isoleucine, and valine (system LIV-I) while the uptake of histidine and proline was unchanged . A similar increase was observed for branched-chain amino-acid binding protein activity . The temperature change did not alter the transport activity for any of these substrates or the level of the binding proteins in an isogenic strain with a normal leucyl-tRNA synthetase . The increase in transport activity observed in the mutant was prevented by inhibitors of protein and RNA synthesis and probably represents an increase in the differential rate of synthesis of the protein(s) required for transport . These experiments demonstrate that the repression of branched-chain amino-acid transport involves the interaction of leucine with its aminoacyl-tRNA synthetase and its cognate leucyl-tRNA species. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3888 - 92 Neutron scattering measurements of separation and shape of proteins in 30S ribosomal subunit of Escherichia coli: S2-S5, S5-S8, S3-S7; Engelman DM et al.; Neutron scattering measurements done on E . coli 30S ribosomal subunit specimens in which specific pairs of proteins were deuterated have enabled us to estimate the distances between the labeled proteins . The distances between centers of gravity of three protein pairs have been determined: S2-S5 (105 A), S3-S7 (115 A), and S5-S8 (35 A) . A method for extracting shape information about these proteins from the neutron scattering profiles is demonstrated . The method shows that S5 and S8 are compact and S2 is extended. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3863 - 7 DNA replication in vitro starting with an intact phiX174 phage; Jazwinski SM et al.; Conversion of the single-stranded DNA in the intact phiX174 phage particle to the duplex replicative form (RF) has been demonstrated in lysates form phage-sensitive cells . The conversion is resistant to rifampicin and requires participation of both a "membrane" fraction of the lysate and a multienzyme replicative system . The lipopolysaccharide phage receptor, while essential, does not replace the membrane fraction . Clear, nonsedimentable extract fractions prepared with a certain nonionic detergent can replace the membrane fraction . Purification of the activity in these extracts by adsorption to polypropylene film yields a fraction with a 5-fold increase in activity relative to lipopolysaccharide and 50-fold increase relative to protein . The low buoyant density (1.03 g/cm3) suggests a high phospholipid or detergent content in this fraction. J Gen Microbiol, 1975 Oct, 90(2), 303 - 10 R factor tetracycline and chloramphenicol resistance in Escherichia coli K12 cmlB mutants; Foster TJ; The isolation of Escherichia coli chromosomal mutants that increased the level of resistance of a partially tetracycline-sensitive mutant of RI00-I is described . Plasmid-less derivatives of these moderately resistant mutants were phenotypically similar to the cmlB mutants described by Reeve (1966, 1968), and also mapped in the same region . The level of intrinsic resistance to both chloramphenicol and tetracycline was increased about twofold . Also, the levels of R factor-determined resistance to these drugs were increased by this host mutation and tetracycline resistance was expressed constitutively . A cmlB accumulated tetracycline at a threefold lower rate than the wild-type strain, and it is proposed that the mutants have an altered permeability to the drugs and that this acts synergistically with the products of the R factor chloramphenicol and tetracycline resistance genes. Infect Immun, 1975 Oct, 12(4), 866 - 80 Adherence of enteropathogenic Escherichia coli to intestinal epithelium in vivo; Hohmann A et al.; Two porcine strains of enteropathogenic Escherichia coli, one possessing K88 antigen and one lacking K88, were orally inoculated into conventional neonatal piglets . Athough both strains caused severe diarrhea, only the K88-possessing strain was able to proliferate in the anterior small intestine . Both K88-possessing and K88-lacking strains were found in large numbers in the posterior small intestine and, using fluorescent antibodies and scanning and transmission electron microscopy, were found adhering to the epitheial surface in these regions . The presence of an unusual surface structure on the bacterial cell of the K88-lacking strain was described. Can J Microbiol, 1975 Oct, 21(10), 1595 - 601 Isolation and partial characterization of an Escherichia coli mutant resistant to colicin A; Lavoie M et al.; An Escherichia coli K12 mutant resistant to colicin A-CA31 apparently through loss of its receptor sites has been isolated and partially characterized . Resistance to colicin A was accompanied with a decreased sensitivity to colicins L-398 and E2-CA42, and to acridine dyes . The mutant strain displayed the same general pattern of tolerance or sensitivity as the parent strain towards eight antibiotics, colicins C, D, E1, E3, F2, F3, G, I, K, and N; phages T1, T2, T5, T6, T7, F2, lambda vir, P1kc, phi 80, and BF23; and to methylene blue, triphenyltetrazolium chloride, ethylenediaminetetraacetate (EDTA), deoxycholate, and sodium dodecyl sulfate . Conjugation and transduction experiments showed that a locus controlling resistance to colicin A-CA31 mapped at 21 min on the genetic map of this E . coli K12 strain. Br J Surg, 1975 Oct, 62(10), 777 - 80 Septic shock; Ledingham IM; The problems associated with septic shock are highlighted, including those which arose during a 3-year prospective study . Mortality in the first year of the study was 71 per cent, falling to 38 per cent in the third year . The reasons for this fall are detailed . The pattern of survival times changed over the 3 years, giving rise to the longer term complications such as stress bleeding, multiple organ failure and superadded infection. Nucleic Acids Res, 1975 Oct, 2(10), 1911 - 29 Hybridization of labeled RNA to DNA in agarose gels; Shinnick TM et al.; Specific DNA restriction endonuclease fragments can be identified after electrophoresis in agarose gels by hybridization in the gel (in situ) to radioactive homologous RNA . RNA-DNA hybrids are detected by autoradiography of the gel . Comparison of band patterns of the autoradiogram and the ethidium bromide stained gel allows the identification of the DNA fragment which is complementary to the RNA probe . The technique is rapid, easy and inexpensive . It is sensitive enough to detect individual genes in a mixture of fragments produced by restriction enzyme digestion of complex cellular DNA . We have used this technique to determine which of the Hin III and Eco R1 fragments of phi80d3ilv+su+7 and E . coli DNAs contain the 5S, 16S and 23S ribosomal RNA (rRNA) genes of E . coli. Nucleic Acids Res, 1975 Oct, 2(10), 1867 - 88 Evidence for tertiary structural RNA-RNA interactions within the protein S4 binding site at the 5'-end of 16S ribosomal RNA of Escherichia coli.+; Ungewickell E et al.; Evidence is presented for tertiary structural interaction(s) (interactions(s) between two regions of an RNA molecule that are widely separated in the RNA sequence) within the 5'-one third of the 16S ribosomal RNA of Escherichia coli that constitutes the binding site of protein S4 . The two main interacting RNA regions were separated by about 120 nucleotides (sections Q to M) of the 16S RNA sequence . A second, smaller gap, of 13 nucleotides, occurred within section C" . The two main interacting regions contain about 150 nucleotides (sections H" to Q) and 160 nucleotides (sections M to C") . They are folded back on one another and, especially in the presence of protein S4, are strongly protected against ribonuclease digestion . The intermediate region (sections Q to M), however, is relatively accessible to ribonucleases in the S4-RNP . By partial removal of subfragments from the RNA complex it was possible to localise the two main interacting sites within sections H" - H and sections I" - C" . Three main criteria for the specificity of the RNA-RNA interactions were invoked and satisfied . The possibility of other tertiary structural RNA-RNA interactions occurring in other regions of the 16S RNA is discussed . Finally, all the structural information on the S4-RNP is summarised and a tentative model is proposed. Nucleic Acids Res, 1975 Oct, 2(10), 1821 - 37 Physical studies of the interaction between the Escherichia coli DNA binding protein and nucleic acids; Molineux IJ et al.; The interaction of nucleic acid with the Escherichia coli DNA-binding protein has been studied by fluorescence emission spectroscopy and sedimentation velocity analysis . The protein binds to single-strand DNA with an apparent equilibrium dissociation constant of 2 X 10(-9) . It binds to the homopolymers poly (dA) and poly (dT) slightly more tightly, but has a larger apparent equilibrium dissociation constant to poly (dC) . The protein also binds tightly to ribohomopolymers and to tRNA, but not to duplex DNA . By the use of defined-length oligonucleotides, it has been shown that the protein binds to DNA in a highly cooperative manner . The extent of cooperativity is seen as the difference in binding between an isolated monomeric protein molecule bound to DNA and two or more molecules binding to contiguous sites. Immunology, 1975 Oct, 29(4), 697 - 707 Effects of mitogens for mouse B lymphocytes on chicken lymphoid cells; Tufveson G et al.; Lipopolysaccharide from E . coli (LPS) and purified protein derivative from tuberculin (PPD) increased the {3H}thymidine incorporation of chicken spleen cells in culture . No such stimulation was observed with dextran sulphate . Thymus and bursa lymphocytes were not stimulated by any of these compounds . Spleen cells from chickens chemically bursectomized with cyclophosphamide treatment showed decreased responses to LPS and PPD, but responded normally to the T-cell mitogen concanavalin A . None of the tested mitogens induced polyclonal antibody synthesis or directly enhanced the primary antibody response to sheep erythrocytes (SRBC) in spleen cell cultures . LPS-coated SRBC, however, enhanced the in vitro antibody response to SRBC . The results suggest a moderate proliferative response of chicken lymphocytes to LPS and PPD, possibly involving B cells, but not further effects comparable to those on mouse B lymphocytes. Eur J Biochem, 1975 Oct 1, 58(1), 59 - 64 Initiation specificity of the poly(cytidylic acid)-dependent Qbeta replicase activity; Feix G et al.; The initiation specificity of RNA synthesis catalysed by the poly(C)-dependent Qbeta replicase activity was investigated with various synthetic ribopolymers as template . It was found that the initiation efficiency of a series of oligo(C) with various chain lengths is proportional to the template size . Synthetic riboheteropolymers containing cytidylic acid were accepted as templates only if they contained at their 3' end a cytidylic acid sequence of more than 5 nucleotides . Such an oligocytidylate sequence served also as an initiator sequence for copying the non-cytidylic-acid-containing part of the heterotemplate . RNA synthesis always began with the incorporation of GTP, even if the 3'-terminating nucleotide of the template was not cytidylic acid. Cell, 1975 Oct, 6(2), 231 - 44 Studies of mouse mitochondrial DNA in Escherichia coli: structure and function of the eucaryotic-procaryotic chimeric plasmids; Chang AC et al.; The mouse mitochondrial DNA genome has been cloned in Escherichia coli by linking it to the pSC101 plasmid replicon at cohesive-ended cleavage sites generated by Eco Rl restriction endonuclease . The four possible configurations of chimeric molecules that contain the nucleotide sequences of mitochondrial DNA in their native relationship were distinguished by Hind III restriction endonuclease digestion and electron microscopic heteroduplex analysis . Chimeric molecules utilize the pSC101 replication origin and do not maintain the "D-loop" region or the low frequency of ribonucleotides found in native mitochondrial DNA . Hybridization of the RNA synthesized in E . coli minicells carrying the four types of chimeras indicates that transcription occurs predominately on the light strand of the mitochondrial DNA in all cases . This result implies that initiation of RNA synthesis occurs within the mitochondrial DNA segment . Although specific polypeptide synthetis is directed by the mitochondrial DNA segment of each of the chimeras in E . coli minicells, the molecular weight distribution of the polypeptides synthesized consists primarily of low molecular weight species and thus differs from that observed in mitochondria in mouse L cells. Biochim Biophys Acta, 1975 Oct 1, 407(2), 240 - 8 Aminoacylation of Phaseolus vulgaris cytoplasmic, chloroplastic and mitochondrial tRNAsMet and of Escherichia coli tRNAsMet by homologous and heterologous enzymes; Gillemaut P et al.; Met-tRNA synthetase from Paseolus vulgaris cytoplasm could be separated from its chloroplastic or mitochondrial counterpart by DEAE-cellulose chromatography, but the Met-tRNA synthetase from the two latter organelles could not be distinguished using DEAE-cellulose, hydroxyapatite or CM-Sephadex chromatography . As revealed by reverse-phase chromatography, bean cytoplasm contains 2 tRNAsMet; only one is charged by chloroplast, mitochondrial or Escherichia coli Met-tRNA synthetase . Mitochondria contain, in addition to the 2 cytoplasmic tRNAsMet, 3 mitochondria-spedific tRNAsMet; 2 can be formylated by the mitochondrial or the E . coli transformylase; all 3 are charged by mitochondrial, chloroplastic or E, coli Met-tRNA synthetase; none is charged by the cytoplasmic enzyme . Chloroplasts contain, in addition to the 2 cytoplasmic tRNAsMet, 3 chloroplast-specific tRNAsMet, different from the mitochondrial tRNAsMet; one is formylatable by the chloroplastic or the E . coli transformylase; all 3 are charged by chloroplastic, mitochondrial or E . coli Met-tRNA synthetase; only one is charged by the cytoplasmic enzyme . Of the 3 E . coli tRNAsMet, only the formylatable species can be charged by bean cytoplasmic, chloroplastic or mitochondrial Met-tRNA synthetase. J Virol, 1975 Oct, 16(4), 838 - 43 Degradation of the viral strand of phiX174 parental replicative-form DNA in a rep- host; Bowman KL et al.; A progressive degradation of the parental viral strand label is observed upon infection of a Rep- mutant of Escherichia coli by 32P-labeled phiX174 . Very little parental label remains in the RF (replicative form) by 47 min after infection . Concomitant with this degradation, replicative intermediates are formed which sediment at 21s, the rate of RF I (supercoiled-closed circular DNA), in a neutral sucrose gradient but which denature and sediment in alkaline gradients as single strands of unit size and larger . These denaturable 21s replicative intermediates have been shown previously to be RF molecules containing an elongated viral strand . Addition of chloramphenicol at 7 min after infection at 30 mug/ml, a concentration sufficient to block RF leads to SS (single strand) synthesis but not RF leads to RF synthesis in a wild-type host cell, reduced the amount of viral strand elongation but did not prevent viral strand degradation . The addition of chloramphenicol at 150 mug/ml at 7 min after infection totally prevents both the degradation of the parental label and the formation of the replicative intermediates with elongated tails . We infer that degradation of the viral strand requires the gene A-mediated nicking of the viral strand but not the concomitant elongation of the viral strand. J Virol, 1975 Oct, 16(4), 1090 - 3 Fate of MS2 proteins synthesized in Escherichia coli exposed to amino acid analogues; Prouty WF; Incorporation of an analoque into MS2 coded proteins prevents the maturation of phages . In addition, there is an alteration in the relative amount of coat protein to replicase protein synthesized, which supports the hypothesis that normal coat protein serves a physiological role as a translation repressor . Further, abnormal proteins, synthesized from the phage genome, are degraded, presumably by a host catabolic system, more rapidly than the normal gene products. J Bacteriol, 1975 Oct, 124(1), 99 - 111 Physiological and biochemical studies on the function of 5-methyluridine in the transfer ribonucleic acid of Escherichia coli; Bjork GR et al.; Matched pairs of transductant strains differing by the presence of absence of 5-methyluridine (ribothymidine) (m5U) in their transfer ribonucleic acid (tRNA) were used to study the function of this modified nucleoside in Escherichia coli . Ordinary measurements of growth rate in different media revealed no effect of the loss of m5U in tRNA . A gene located close to trmA (the structural cistron for the methyltransferase that produces m5U in tRNA), however, was found to reduce the growth rates significantly, depending on the medium and the temperature of cultivation . Measurement of codon recognition, macromolecular composition, tRNA binding to the ribosome, and the rate of protein chain elongation in vivo indicated no disadvantage caused by the lack of m5U . The regulation of ilv and his operons seemed also to be unaffected by the absence of m5U in the tRNA . In a mixed population experiment, however, cells possessing m5U in their tRNA seemed to have a distinct advantage over cells lacking this modified nucleoside . This experiment provides the first indication of the overall value of m5U in tRNA. J Bacteriol, 1975 Oct, 124(1), 92 - 8 Transductional mapping of gene trmA responsible for the production of 5-methyluridine in transfer ribonucleic acid of Escherichia coli; Bjork GR; The gene trmA, responsible for the production of 5-methyluridine (ribothymidine) in transfer ribonucleic acid, has been located at 79 min on the chromosomal map of Escherichia coli K-12 . In five-factor crosses the gene order was shown to be argH-trmA-rif-thiA-metA . The co-transduction frequency between argH and trmA was 65% . Furthermore, the trmA5 mutation was shown to be recessive, in agreement with the notion that the trmA gene is the structural gene for the transfer tibonucleic acid (5-methyluridine) methyltransferase. J Bacteriol, 1975 Oct, 124(1), 435 - 44 Lag in adaptation to lactose as a probe to the timing of permease incorporation into the cell membrane; Koch AL; If bacteria are incapable of forming and incorporating proteins into the cytoplasmic membranes in all phases of the cell cycle, then not all cells from an asynchronous culture should be capable of growth when switched to a new carbon and energy source whose metabolism requires new membrane function . The transfer of an inducible culture to low lactose provides such a situation since the cells cannot grow unless galactoside permease can function to concentrate the lactose internally . From such experiments, it was concluded that the Y gene product of the lac operon is synthesized, incorporated, and can start functioning in active transport, at any time throughout the bulk of the cell cycle . Not only were the lags before growth re-ensued much shorter than would be expected if the membrane transport capability could only be developed in a small portion of the cycle, but brief pulses of a gratuitous inducer shortened the lags much further . Three types of Escherichia coli ML 30 culture were studied: cells that had exhausted the limiting glucose; cells taken directly from glucose-limited chemostats; and a washed suspension of highly catabolite repressed cells from cultures grown in high levels of glucose and gluconate . The growth studies reported here were performed on-line with a minicomputer . They represent at least an order of magnitude increase in accuracy in estimating growth parameters over previous instrumentation. J Bacteriol, 1975 Oct, 124(1), 26 - 38 Mutations affecting transport of the hexitols D-mannitol, D-glucitol, and galactitol in Escherichia coli K-12: isolation and mapping; Lengeler J; Mutants of Escherichia coli K-12 unable to grow on any of the three naturally occurring hexitols D-manitol, D-glucitol, and galactitol and, among these specifically, mutants with altered transport and phosphorylating activity have been isolated . Different isolation procedures have been utilized, including suicide by D-{3H}mannitol, chemotaxis, and resistance to the toxic hexitol analogue 2-deoxy-arabino-hexitol . Mutations thus obtained have been mapped in four distinct operons . (i) Mutations affecting an enzyme II-complexmt1 activity of the phosphoenolpyruvate-dependent phosphotransferase system all map in gene mtlA . This gene has previously been shown (Solomon and Lin, 1972) to be part of an operon, mtl, located at 71 min on the E . coli linkage map containing, in addition to mtlA, the cis-dominant regulatory gene mtlC and mtlD, the structural gene for the enzyme D-mannitol-1-phosphate dehydrogenase . The gene order in this operon, induced by D-mannitol, is mtlC A D . (ii) Mutations in gene gutA affecting a second enzyme II-complexgut of the phosphotransferase system map at 51 min, clustered in operon gutC A D together with the cis-dominant regulatory gene gutC and the structural gene gutD for the enzyme D-glucitol-6-phosphate dehydrogenase . The gut operon, previously called sbl or srl, is induced by D-glucitol . (iii) Mutations affecting the transport and catabolism of galactitol are clustered in a third operon, gatC A D, located at 40.5 min . This operon again contains a cis-dominant regulatory gene, gatC, the structural gene gatD for galactitol-1-phosphate dehydrogenase, and gene gatA coding for a thrid hexitol-specific enzyme II-complexgat . Other genes coding for two additional enzymes involved in galactitol catabolism apparently are not linked to gatC A D . (iv) A fourth class of mutants pleiotropically negative for hexitol growth and transport maps in the pts operon . Triple-negative mutants (mtlA gutA gatA) do not have further transport or phosphorylating activity for any of the three hexitols. J Bacteriol, 1975 Oct, 124(1), 140 - 8 Isolation of an Escherichia coli mutant deficient in glutathione synthesis; Fuchs JA et al.; A mutant of Escherichia coli that contains essentially no detectable glutathione has been isolated . The mutant contains a very low level of the enzyme glutathione synthetase and accumulates lambda-glutamyl cysteine at a concentration approximately equal to the level of glutathione found in its parent . No significant differences in growth were observed between the mutant and its parent . However, the activity of at least one enzyme was found to be affected by the absence of glutathione; the specific activity of the B1 subunit of ribonucleoside diphosphate reductase was greatly reduced . The possibility that the decreased B1 activity is due to a mutation in the structural gene coding for B1 or its regulatory gene could be eliminated . This suggests that one role of glutathione in the cell is to maintain at least this one protein in an active state . We propose the designation gshB for the gene coding for glutathione synthetase. Clin Pediatr (Phila), 1975 Oct, 14(10), 934 - 8 Cephalexin in the management of bacteriuria; Fennell RS 3rd et al.; This report evaluates the effectiveness of cephalexin in the treatment of bacteriuria in 93 children . Cephalexin therapy eradicated sensitive organisms in 97 per cent of the cases regardless of recurrence, structural abnormality, or status of renal function . Nevertheless, recurrences with a new, resistant organism occurred significantly in certain patients, expecially those with major anatomic abnormalities, during a six-week follow-up period . The incidence of drug reactions was low. Can J Comp Med, 1975 Oct, 39(4), 421 - 6 Polyserositis in pigs due to generalized Escherichia coli infection; Nielsen NC et al.; A fibrinous polyserositis syndrome due to generalized Escherichia coli infection in pigs was observed in 13 out of 17 systematically monitored herds . The mortality rate was approximately 0.1% among liveborn pigs . The occurrence was usually sporadic but a minor enzootic was observed in one herd . Most of the affected pigs succumbed during first or second week of life but cases were observed throughout the suckling period . The clinical signs included marked depression, anorexia, rough haircoat, laboured respiration and death in two to five days . Predominant gross pathological lesions were signs of septicaemia and a voluminous, gelatinous and fibrinous exudate in the pleural, the pericardial and the peritoneal cavities . Frequently also a firbinous polyarthritis and a fibrinous pneumonia were present . The majority of the isolated invasive E . coli strains were nonhaemolytic . Serologically 11 different E . coli O groups were encountered . O group most frequently represented was 020 . None of the examined E . coli strains belonged to the serogroups which frequently are associated with enteropathogenicity in pigs. Can J Comp Med, 1975 Oct, 39(4), 371 - 6 Relationships of intestinal enzymes and serum antitoxin to the pig response to Escherichia coli enterotoxin; Lariviere S et al.; The intestinal loop technique was used to evaluate the response of three week old piglets to the heat labile (LT) and the heat stable (ST) enterotoxins produced by Escherichia coli F11(P155) . The serum anti-LT activity and the lipase, amylase and trypsin activities in the jejunal lumen of these pigs were determined . Piglets responded independently ti each toxin and no relationship between these responses and serum anti-LT activity or the enzyme activities of the jejunal content could be demonstrated. J Clin Invest, 1975 Oct, 56(4), 870 - 9 Behavior of eosinophil leukocytes in acute inflammation . II . Eosinophil dynamics during acute inflammation; Bass DA; The marked diminution in the number of circulating eosinophils, which has been shown to occur during acute bacterial infections, is a distinctive aspect of eosinophil physiology and of the host response to acute infection . The mouse rendered eosinophilic by infection with trichinosis provides a suitable model for study of the eosinopenic response induced by acute inflammation . The alterations in eosinophil dynamics associated with acute inflammatory reactions in trichinous mice were studied with pneumococcal abscesses, with Escherichia coli pyelonephritis, with Coxsackie viral pancreatitis, and with acute subcutaneous inflammation due to turpentine . Each of these stimuli of acute inflammation markedly suppressed the eosinophilia of trichinosis . This suggests that the eosinopenia is a response to the acute inflammatory process rather than the response to a specific type of pathogen . These studies apply quantitative techniques to ascertain the effects of acute inflammation on eosinophil production in bone marrow and on distribution of eosinophils in the peripheral tissues . From these observations, it is apparent that the initial response to acute inflammation includes a rapid drop in numbers of circulating eosinophils, a rapid accumulation of eosinophils at the periphery of the inflammatory site, and an inhibition of egress of eosinophils from the bone marrow . With prolongation of the inflammatory process, inhibition of eosinopoiesis occurs. J Bacteriol, 1975 Oct, 124(1), 470 - 5 Localization of D-lactate dehydrogenase in membrane vesicles prepared by using a french press or ethylenediaminetetraacetate-lysozyme from Escherichia coli; Futai M et al.; The localization of D-lactate dehydrogenase in membrane vesicles prepared from Escherichia coli was studied using antibody against the purified enzyme . The activity of D-lactate dehydrogenase and D-lactate-dependent oxygen uptake of membrane vesicles prepared by using a French press were completely inhibited by this antibody, suggesting that the enzyme is localized on the outside of these vesicles . This and previous results (Futai, 1974) strongly indicate the inversion of these vesicles . The D-lactate dehydrogenase and D-lactate-dependent oxygen uptake of membrane vesicles prepared by treatment with ethylenediaminetetraacetate-lysozyme were inhibited about 15% by the antibody, whereas proline transport of the vesicles was insensitive to antibody . These results suggest that most of the membrane vesicles have D-lactate dehydrogenase on the inside of the membrane and that such vesicles transport amino acids . This essentially confirms the results of Short, Kaback, and Kohn (1975) . However, unlike them we observed that a small but significant portion of activity was sensitive to the antibody as shown above . This portion may represent the completely inverted vesicles in the preparation . Ferricyanide reductase activity cannot be detected in spheroplasts, but about 30 to 50% of the total was detected in membrane vesicles prepared by treatment with ethylenediaminetetraacetate . This confirms our previous findings with membrane prepared by a slightly different procedure . It is concluded that in these vesicles about half the reactive sites for ferricyanide are moved from inside to outside the membrane, whereas 85% of the D-lactate dehydrogenase remains inside the membrane. Jpn J Microbiol, 1975 Oct, 19(5), 373 - 80 Isolation and properties of Escherichia coli mutants which are nonpermissive for the growth of phage lambda; Takahashi S et al.; By selecting survivors of lambda phage infection, mutants of Escherichia coli K12 that block reproduction cycle of the phage have been isolated . Fourteen of these phage-tolerant mutants (lam mutants) were chosen and characterized biochemically and genetically . It was shown that these mutants were tolerant to infection by all the lambdoid phages, except for few cases, but they were susceptible to infection by a non-lambdoid temperate phage (phi299), P1 or T phages . The mutants can be divided into at least three groups: (1) A mutant (lam 16) strain that seems to block normal penetration of phage DNA: (2) Three mutant (lam 64, lam 67 and lam 71) strains that block an "early" step(s) of phage growth, including phage DNA synthesis: (3) Six mutant (lam 24, lam 25, lam 26, lam 27, lam 646 and lam 6) strains that block normal functioning of the gene E products and produce unusual head structures . Some lambdoid phages and lambda mutants that overcome the interference by the lam mutations have been obtained, and were used as tools for characterizing the host mutations . Two (lam 12 and lam 13) mutant strains and one (lam 1) mutant were inferred as affecting the expression of "late" genes, and early gene, respectively, by this test. Jpn J Microbiol, 1975 Oct, 19(5), 363 - 71 Mutants of Escherichia coli which block head formation of lambda; Hidaka S et al.; Five mutants of Escherichia coli K12 (lam 24, lam 25, lam 26, lam 27 and lam 646) that block head formation of lambda are described . In vitro complementation tests and electron microscopy demonstrated that in these bacteria phage tails were produced normally, whereas head formation was abnormal, aberrant head-related structures being produced . In lysates prepared from lam 24, lam 25 and lam 26, monsters and empty heads without tail were the predominant structures, whereas in lysates from lam 27 and lam 646, petit lambda and empty heads were the most common structures . The five lam mutations were located in two regions on the bacterial chromosome; lam 24, lam 25 and lam 26 were near the dnaB gene and lam 27 and lam 27 and lam 646 near the lac gene . It was suggested that the former three mutants are new isolates that belong to GroE mutants, whereas the latter two comprise a new group of mutants . Analyses of phage mutants (ov mutants) that overcome the interference by the lam 646 mutation revealed that this mutation blocks normal expression of the gene E of lambda. Jpn J Microbiol, 1975 Oct, 19(5), 349 - 54 Induction of phage production in the lysogenic Escherichia coli by hydroxyurea; Shimada K et al.; 1) Hydroxyurea, a reversible DNA synthesis inhibitor, was used to study the mechanism of prophage lambda induction in Escherichia coli K12 . Induction of prophage was judged on two criteria: increase of phage-producing cells and loss of colony-forming ability of the cells . 2) Hydroxyurea induced an increase of phage-producing cells only in lysogenic strains known to be inducible with ultraviolet irradiation for prophage development and not in strains such as E . coli K12 (lambdaind-) or E . coli K12 recA (lambda+) . 3) When protein synthesis was inhibited, hydroxyurea did not increase phage-producing cells of lysogenic strains; it showed a bacteriocidal effect on lysogenic recA+ strains, but not on nonlysogenic strains . 4) The sensitivity of E . coli K12 recA to hydroxyurea was independent of whether or not the cells were lysogenic . 5) From the results it is suggested that certain steps leading to loss of colony-forming ability (i.e . prophage induction) do not require de novo protein synthesis but require the presence of the host recA+ gene. Mol Biol Rep, 1975 Oct, 2(3), 233 - 9 A directed alteration of ribonuclease specificity . Hydrolysis of polyribonucleotides containing modified cytosine bases; Mazo AM et al.; The action of ribonucleases on poly and oligoribonucleotides containing cytosine bases modified by methoxyamine and bisulphite was examined . Resistance of phosphodiester bonds in (Cp)nXp (where n greater than or equal to 1 and X stands for A, G or U) to T2 RNase hydrolysis was observed if substrates were modified chemically . The phenomenon formed the basis for isolation of (Cp)nXp blocks as an additional tool in sequence investigations . After modification of cytosine pancreatic RNase was unable to hydrolyse (Cp)nUp blocks . Therefore the specificity of pyrimidyl RNase may be narrowed to 'uridyl RNase'. J Gen Microbiol, 1975 Oct, 90(2), 321 - 35 Genes involved in the uptake and catabolism of gluconate by Escherichia coli; Bachi B et al.; The isolation and properties of a mutant of Escherichia coli K12 that is totally unable to take up and utilize gluconate are described . Genetical analysis shows this phenotype to be associated with two lesions . One phenotype, designated GntM-, is the result of a mutation in a gene co-transducible with malA; the other, designated GNTS-, is the result of a mutation in a gene (GntS) co-transducible with fdp . The GntS--phenotype differs little from that of wild-type cells, but GntM- GntS+ organisms grow on gluconate only after a prolonged lag and form a gluconate uptake system that is strongly repressed by pyruvate . Moreover, such GntM- mutants readily give rise to further mutants that form a gluconate uptake system, gluconate kinase and 6-phosphogluconate dehydratase consititutively; in partial diploids, this constitutivity is recessive to the inducible character . It is postulated that the GntM- phenotype is due to malfunction of a negative control gene gntR, and that gntS+ specifies the activity of a gluconate uptake system. Nucleic Acids Res, 1975 Oct, 2(10), 1851 - 65 Sequences spanning the EcoRI substrate site; Garfin DE et al.; Substrate recognition by the EcoRI restriction endonuclease was investigated by analysis of the nucleotide sequences at the sites of enzymatic cleavage in various DNA molecules . 5'-end labeling and homochromatographic fingerprinting led to the determination of a 17-base-pair sequence spanning the EcoRI site of simian virus 40 DNA and a 15-base-pair sequence overlapping the EcoRI site of Col El plasmid DNA . Three other DNAs were similarly tested, although extended sequences were not determined in these cases . The EcoRI site was shown to be symmetric double-stranded equivalent of -N-G-A-A-T-T-C-N-. J Bacteriol, 1975 Oct, 124(1), 585 - 8 Role for deoxyribonucleic acid ligase in deoxyribonucleic acid polymerase i-dependent repair synthesis in toluene-treated escherichia coli; Billen D et al.; In a toluene-treated mutant of Escherichia coli K-12 having a temperature-sensitive, conditionally lethal mutation in the structural gene for deoxyribonucleic acid (DNA) ligase, an extensive DNA repair synthesis occurred in X-irradiated cells at the nonpermissive temperature, 42 C . At the permissive temperature, 30 C, nearly normal semiconservative synthesis and limited repair synthesis were observed when DNA ligase was activated by the addition of nicotinamide adenine dinucleotide. J Bacteriol, 1975 Oct, 124(1), 380 - 90 Metabolism of cyclic adenosine 3',5'-monophosphate and induction of tryptophanase in Escherichia coli; Botsford JL; The relationship between cyclic adenosine 3',5'-monophosphate (cyclic AMP) metabolism and the induction of tryptophanase and beta-galactosidase was studied in several strains of Escherichia coli grown with succinate, acetate, glycerol, or glucose as the carbon source . No consistent relationship between the intracellular concentration of cyclic AMP in the several strains cultured and the various carbon sources was discerned . In E . coli K-12-1 the induction of tryptophanase was found to vary in the order: succinate greater than acetate greater than glycerol greater than glucose, and that of beta-galactosidase was found in the order: glycerol greater than acetate greater than succinate greater than glucose . Rate of accumulation of cyclic AMP in the culture filtrate was in the order: succinate greater than acetate greater than glycerol greater than glucose . The addition of glycerol to E . coli K-12-1 grown in acetate caused inhibition of tryptophanase and slight inhibition of accumulation of extracellular cyclic AMP . These same conditions caused beta-galactosidase induction to be stimulated . The addition of exogenous cyclic AMP to cultures grown with four different carbon sources had an effect characteristic for each of the two enzymes studied as well as each individual carbon source . The results suggest that there are control elements distinct from cyclic AMP and its receptor protein which respond to the catabolic situation of the cell. J Bacteriol, 1975 Oct, 124(1), 252 - 61 Genetic analysis of succinate utilization in enzyme I mutants of the phosphoenolpyruvate: sugar phosphotransferase system in Escherichia coli; Alexander JK et al.; Studies on the reversion characteristics of Escherichia coli strains carrying various mutations in the pts region have led to the recognition of a mutation, suc-1, with a previously undescribed phenotype . Strains carrying the suc-1 mutation grow normally on most sources of carbon but are unable to utilize succinate effectively . The suc-1 mutation can be separated genetically from the tightly linked ptsI6 mutation . Reversion of suc-1 mutants for growth on succinate yields interesting classes of suppressor mutations. J Bacteriol, 1975 Oct, 124(1), 161 - 6 Operator-promoter functions in the threonine operon of Escherichia coli; Gardner JF et al.; The prophage curing properties of secondary-site lysogens of coliphage lambda have been studied . The site of integration in the original lysogen (L79) is within the ooerator-promoter region of the thr operon . As a result, expression of the thr enzymes is reduced, and the strain is a leaky threonine auxotroph . Heat pulse curing of strain L79 and a thr+ lysogenic revertant (L79-20) showed that heat pulse curing of both lysogens was int and xis dependent and occurred by correct excisions of the prophage . The heat pulse curing restored strain L79 to prototrophy whereas strain L79-20 synthesized the thr enzymes constitutively and at high levels . This indicates that the reversion mutation in strain L79-20 occurred outside of the prophage and within the operator-promoter region of the thr operon . In contrast, spontaneous curing of both lysogens occurred by both correct and incorrect excisions . Spontaneously cured derivatives of strain L79-20 gave rise to three classes of regulatory mutants affecting operator and promoter functions to the thr operon. Eur J Biochem, 1975 Oct 1, 58(1), 87 - 94 Poly(A) synthesis in T2L phage-infected Escherichia coli . A combination of polynucleotide phosphorylase and ATPase; Wunderli W et al.; In crude extracts of T2L phage-infected Escherichia coli cells an enzyme activity was found that produced poly(A) from ATP as substrate . Purification of the extract led to the isolation of two enzymes, a polynucleotide phosphorylase and an ATPase . The polynucleotide phosphorylase possessed the same properties as the well-known enzyme from uninfected cells and its molecular weight was about 265 000 . The ATPase was purified to over 90% purity; its molecular weight was estimated to be about 165 000 with three subunits of 55 000 . The characterization of this enzyme showed that it was different from any ATPase known so far . Mg2+ cannot be replaced by Ca2+, as it can from the membrane-bound ATPases . The only product yielded by the enzyme was ADP; it was very specific for ATP, other ribonucleotide triphosphates being practically unaffected . The rate of ATP splitting was found to be very high, the turnover number being 2.51 X 10(4) min-1 at 37 degrees C . Even at 0 degree C the enzyme was still active . The optimal assay conditions for ATPase turned out to be very similar to those of polynucleotide phosphorylase . Thus the combination of the two enzymes very efficiently produced poly(A) from ATP . In this combination the polynucleotide phosphorylase was the rate-limiting enzyme, since its turnover number was about 40 times lower than that of the ATPase . The evaluation of a variety of properties of the poly(A)-synthesizing constituent found in the crude extracts led us to conclude that this activity arises from the combined action of ATPase and polynucleotide phosphorylase, and is not due to a poly(A) polymerase. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3970 - 4 Ubiquinone-mediated coupling of NADH dehydrogenase to active transport in membrane vesicles from Escherichia coli; Stroobant P et al.; Addition of ubiquinone-1 to E . coli ML 308-225 membrane vesicles dramatically increases coupling between NADH oxidation and active transport such that initial rates and steady-state levels of lactose and amino-acid accumulation are comparable to those observed during D-lactate oxidation . Similar but less dramatic effects are observed with the quinone and succinate or L-lactate . In the presence of NADH and ubiquinone-1, the vesicles also generate a membrane potential (interior negative) that is similar in magnitude to that observed in the presence of D-lactate . Stimulation of NADH-dependent transport by ubiquinone-1 cannot be accounted for by increased rates of oxidation of NADH, and the effect of the quinone on NADH-dependent lactose transport is not observed in vesicles depleted of NADH dehydrogenase activity . Thus, it is apparent that ubiquinone-1 shunts electrons from NADH dehydrogenase {NADH:(acceptor)oxidoreductase; EC 1.6.99.3} to the portion of the respiratory chain containing the energy-coupling site . The findings demonstrate unequivocally that inefficient coupling of NADH oxidation to active transport cannot be due to the presence of inverted vesicles . In addition, they provide further support for specific localization of the energy-coupling site. Proc Natl Acad Sci U S A, 1975 Oct, 72(10), 3907 - 11 phiX174 DNA-dependent DNA synthesis in vitro: requirement for P1 ban protein in dnaB mutant extracts of Escherichia coli; Schuster H et al.; Ammonium sulfate fractionation of crude extracts of E . coli yields a soluble enzyme fraction (about 25-fold purification) that catalyzes the conversion of phiX174 single-stranded DNA to duplex DNA . The reaction is rifampicin-resistant, requires single-stranded DNA, Mg++, deoxynucleoside triphosphates, and ATP, and is stimulated by KCl . Such soluble enzyme fractions were prepared from E . coli strains carrying the prophage mutant P1bac, in which the viral dnaB analog (ban) protein is expressed constitutively, or P1bacban, in which the expression of ban protein is prevented . DNA-synthesizing activity of ban protein containing fractions from wild-type or dnaB(P1bac) lysogens was more temperature-resistant than that from E . coli containing only wild-type dnaB protein, whereas that from dnaB(P1bacban) lysogens of dnaB cells was extremely thermolabile . It is suggested that the temperature-resistant DNA synthesis with fractions from P1bac lysogens is mediated by the P1 ban protein. Mol Biol Rep, 1975 Oct, 2(3), 282 - 8 Identification of the 30 S protein adjacent to peptidyl transferase catalytic center of Escherichia coli ribosomes; Hatayama T et al.; Iodoacetylphenylalanyl-tRNAPhe was used as an affinity label to localize the ribosomal components involved in the peptidyl transferase catalytic center of Escherichia coli ribosomes . When labeling was carried out at pH 5.0, the affinity label could specifically label the ribosomal components which comprise the catalytic center . Analysis of ribosomal proteins which had reacted with the affinity label revealed that a 30 S subunit protein, S 20, was located at or near to the ribosomal binding site of the 3'-terminus of aminoacyl- or peptidyl-tRNA. Mol Gen Genet, 1975 Sep 29, 140(2), 123 - 35 Isolation of phage P2-186 intervarietal hybrids and 186 insertion mutants; Bradley C et al.; Intervarietal hybrids formed between coliphages P2 and 186 have been isolated and their preliminary genetic characterization described . Three insertion mutants of 186 have also been isolated. J Biol Chem, 1975 Sep 25, 250(18), 7195 - 203 Evidence for the coordinate control of glycogen synthesis, glucose utilization, and glycolysis in Escherichia coli . II . Quantitative correlation of the inhibition of glycogen synthesis and the stimulation of glucose utilization by 2,4-dinitrophenol with the effects on the cellular levels of glucose 6-phosphate, fructose, 1,6-diphosphate, and total adenylates; Dietzler DN et al.; In cultures of Escherichia coli W4597(K) and G34 under various nutritional conditions the rates of glucose utilization and cellular levels of fructose-1,6-P2 are quantitatively related by the Hill equation where the value of the Hill coefficient is approximately equal to 2 . This is the first evidence that fructose-P2, or any metabolite which covaries with fructose-P2, modulates glucose utilization in E . coli . In light of previous observations from our laboratory this new observation and those in the succeeding report provide the first evidence that in E . coli glycolsis, glycogen synthesis and glucose utilization are coordinately regulated, thus providing for the coupling of ATP utilization and production under various metabolic circumstances . Alterations in the level of ATP apparently affect the velocity of phosphofructokinase, the rate-limiting enzyme in glycolsis, altering the cellular levels of glucose-6-P or fructose-P2 . Changes in the levels of these hexose phosphates are quantitatively related to alterations in the rates of glucose utilization and glycogen synthesis in the intact E . coli cell. J Biol Chem, 1975 Sep 25, 250(18), 7188 - 93 Evidence for the coordinate control of glycogen synthesis, glucose utilization, and glycolysis in Escherichia coli . I . Quantitative covariance of the rate of glucose utilization and the cellular level of fructose 1,6-diphosphate during exponential growth and nutrient limitation; Dietzler DN et al.; In cultures of Escherichia coli W4597(K) and G34 under various nutritional conditions the rates of glucose utilization and cellular levels of fructose-1,6-P2 are quantitatively related by the Hill equation where the value of the Hill coefficient is approximately equal to 2 . This is the first evidence that fructose-P2, or any metabolite which covaries with fructose-P2, modulates glucose utilization in E . coli . In light of previous observations from our laboratory this new observation and those in the succeeding report provide the first evidence that in E . coli glycolysis, glycogen synthesis and glucose utilization are coordinately regulated, thus providing for the coupling of ATP utilization and production under various metabolic circumstances . Alterations in the level of ATP apparently affect the velocity of phosphofructokinase, the rate-limiting enzyme in glycolysis, altering the cellular levels of glucose-6-P or fructose-P2 . Changes in the levels of these hexose phosphates are quantitatively related to alterations in the rates of glucose utilization and glycogen synthesis in the intact E . coli cell. J Biol Chem, 1975 Sep 25, 250(18), 7099 - 105 Uridine diphosphate galactose-4-epimerase . Uridine monophosphate-dependent reduction by alpha- and beta-D-glucose; Kang UG et al.; Rates of UMP-dependent reduction of the DPN+ associated with Escherichia coli UDP-galactose-4-epimerase at 27 degrees and 0.2 M ionic strength in 0.1 M Tris-HCl buffer, pH 8.5, are reported . The reaction exhibits excellent pseudo-first order behavior when D-glucose is at anomeric equilibrium . The effects of {UMP} and {glucose} on the observed first order rate constants are consistent with the following equation . The symbols phi are empirical parameters . (See article) . The data indicate that the pathway involves random equilibrium binding of UMP and glucose followed by rate-limiting decomposition of the ternary complex to epimerase-DNPH . The binding parameters indicate that the principal activating effect of UMP is not simply to increase the affinity of the enzyme for glucose . UMP appears to increase the reactivity or availability of enzyme-bound DPN+ . The kinetic isotope effect for the reaction of D-}1-2H}glucose (kH/kD) is 4.2, which confirms that C-1 is oxidized and that hydride transfer is rate limiting . Both of the purified anomers, alpha- and beta-D-glucose, reduce the enzyme-bound DPN+ . As indicated by the deviations from pseudo-first order kinetics because of concurrent mutarotation, the beta anomer is the more reactive, reacting about 4 to 5 times faster than the alpha anomer at concentrations well below saturation . Is is suggested that the lack of stereo-specificity in this reaction may be attributed to the two anomers being productively bound with their opposite faces projecting toward C-4 of bound DPN+ . Nonstereospecific oxidation of alpha- and beta-D-glucose may be a model for the mechanism of UDP-hexose epimerization, which also involves nonstereospecific hydride transfer. J Biol Chem, 1975 Sep 25, 250(18), 7307 - 12 A novel oligoribonuclease of Escherichia coli . I . Isolation and properties; Niyogi SK et al.; A new ribonuclease has been isolated from Escherichia coli . The enzyme is present in the 100,000 times g supernatant fraction and has been purified over 200-fold . Studies of the enzyme reveal that: 1 . The enzyme shows a marked preference for oligoribonucleotides; indeed, the reaction rate is inversely proportional to the chain length of the substrate . The enzyme does not attack polynucleotides even at high concentrations of enzyme and has no detectable DNase activity . 2 . The enzyme is stimulated strongly by Mn2+, less strongly by Mg2+, and not at all by Ca2+ and monovalent cations . 3 . The enzyme is purified free of RNase I, RNase II, RNase III, polynucleotide phosphorylase, and other known ribonucleases of E . coli . The enzyme displays identical properties when isolated from mutants of E . coli that are deficient in the above ribonucleases . 4 . The enzyme has a marked thermostability, a point of further distinction from RNase II. J Biol Chem, 1975 Sep 25, 250(18), 7153 - 8 Mutants of Escherichia coli defective in membrane phospholipid synthesis . Phenotypic suppression of sn-glycerol-3-phosphate acyltransferase Km mutants by loss of feedback inhibition of the biosynthetic sn-glycerol-3-phosphate dehydrogenase; Bell RM et al.; Revertants of Escherichia coli mutants defective in the first enzyme of membrane phospholipid synthesis, sn-glycerol-3-phosphate (glycerol-P) acyltransferase, were investigated . These glycerol-P acyltransferase mutants, selected as glycerol-P auxotrophs, contained membranous glycerol-P acyltransferase activity with an apparent Km for glycerol-P 10 times higher than the parental activity . The glycerol-P acyltransferase activity was also more thermolabile in vitro than the parental activity . Most revertants no longer requiring glycerol-P for growth regained glycerol-P acyltransferase activity of normal thermolability and apparent Km for glycerol-P . However, two novel revertants were isolated which retained an abnormal glycerol-P acyltransferase activity . The glycerol-P dehydrogenase activities of these novel revertants were about 20-fold less sensitive to feedback inhibition by glycerol-P . The feedback-resistant glycerol-P dehydrogenase co-transduced with gpsA, the structural gene for the glycerol-P dehydrogenase . Further transduction experiments demonstrated that the feedback resistant glycerol-P dehydrogenase phenotypically suppressed the glycerol-P acyltransferase Km lesion . The existence of the class of glycerol-P auxotrophs which owe their phenotype to the glycerol-P acyltransferase Km lesion therefore depends on the feedback regulation of glycerol-P synthesis in E . coli. J Biol Chem, 1975 Sep 25, 250(18), 7147 - 52 Mutants of Escherichia coli defective in membrane phospholipid synthesis . Properties of wild type and Km defective sn-glycerol-3-phosphate acyltransferase activities; Bell RM; The sn-glycerol-3-phosphate (glycerol-P) acyltransferase, the first enzyme of membrane phospholipid synthesis in Escherichia coli, was investigated in a wild type and a mutant strain defective in this activity . The mutant strain, selected as a glycerol-P auxotroph, was previously shown to contain a glycerol-P acyltransferase activity with an apparent Km for glycerol-P 10 times higher than that of its parent or revertants . The membranous mutant glycerol-P acyltransferase but did not appear to be thermolabile in vivo . Revertants no longer requiring glycerol-P for growth, showed glycerol-P acyltransferase activity with thermolability properties similar to the wild type . The second phospholipid biosynthetic enzyme, 1-acylglycerol-P acyltransferase, was not thermolabile in membranes containing a thermolabile glycerol-P acyltransferase activity . The pH optimum for the mutant acyltransferase was over 1 pH unit higher than that of the parental activity . Further, the mutant and wild type glycerol-P acyltransferase differed in their response to magnesium chloride and potassium chloride . The palmitoyl-CoA dependence of the wild type and mutant glycerol-P acyltransferase activities were different . The mutant glycerol-P acyltransferase activity was inhibited greater than 90% by Triton X-100 under conditions where the wild type activity was not affected . These experiments provide novel information about the wild type glycerol-P acyltransferase activity of E . coli and provide six additional lines of evidence for the mutant character of the glycerol-P acyltransferase in the mutant strains. J Biol Chem, 1975 Sep 25, 250(18), 7313 - 9 A novel oligoribonuclease of Escherichia coli . II . Mechanism of action; Datta AK et al.; Detailed studies of the mechanism of action of the novel oligoribonuclease of Escherichia coli described in the previous paper (1) led to the following conclusions . 1 . The enzyme prefers a free 3'-hydroxyl group for its action . 2 . The enzyme attacks the oligoribonucleotide substrate in a sequential manner from the 3' end producing 5'-ribonucleotides . 3 . The mode of attack appears to be processive; the enzyme acts by degrading one oligoribonucleotide chain to completion before proceeding to the hydrolysis of another chain . 4 . The reaction rate is inversely proportional to the chain length of the substrate; however, the enzyme has a higher affinity for longer chains . 5 . The enzyme activity is markedly inhibited by secondary structure; oligoribonucleotides combined with complementary polyribonucleotides are attacked poorly below the melting temperature of the complex and efficiently above the melting temperature . 6 . The enzyme is inhibited by 5'-nucleotides of adenine and guanine; those of cytosine and uracil have a much smaller effect . The enzyme is not inhibited by 3'-nucleotides . 7 . Studies with dinucleoside monophosphate show highest reaction rates with pyrimidine sequences in the order: CpCgreater than UpUgreater than CpUgreater than UpC . The presence of guanine at the 3' end is strongly inhibitory, and reaction rates are CpGgreater than UpG=ApGgreater than GpG. Biochemistry, 1975 Sep 23, 14(19), 4261 - 6 The effect of Mn2+ and Co2+ on the activities of a zinc metallodipeptidase from a mouse ascites tumor; Patterson EK et al.; Kinetic studies of the effect of addition of Co2+ or Mn2+ to a highly purified dipeptidase from Ehrlich-Lettre mouse ascites tumor cells show that these metals specifically activate the hydrolyses of certain classes of dipeptides . This enzyme was previously (S . Hayman and E . K . Patterson, 1971, J . Biol . Chem . 246, 660) reported to be a Zn-metalloenzyme . It is now shown that Zn is the only metal that can partially restore the activity of the EDTA-inhibited dipeptidase in cleaving Ala-Gly . Addition of Co2+ increases the Vmax of N-terminal Gly-dipeptides with increase in Km while addition of Mn2+ primarily activates the hydrolysis of Pro-Gly, again with increases in both Vmax and Km . Prior incubation (5 min, 30 degrees) of the dipeptidase with the appropriate metal ions causes decrease in initial lag time in the Co2+-activated hydrolysis of Gly-Gly and the Mn2+-activated hydrolysis of Pro-Gly . Long-term (6-19 hr, 0 degrees) incubation of the enzyme with Co2+ results in loss of activity toward Ala-Gly with a concomitant 13-fold increase in the rate of Gly-Gly hydrolysis and loss of 70% of the Zn2+ from the dipeptidase; these effects can be partially reversed by addition of Zn2+ . In contrast, long-term incubation of the enzyme with Mn2+ results in no loss of Zn2+ and a twofold increase in activity toward Pro-Gly . One affinity constant of 1.4 muM for Co2+ and two constants of 0.23 and 27 muM for Mn2+ were determined by kinetic experiments . Comparison of the properties of this tumor enzyme with a dipeptidase purified in our laboratory from Escherichia coli B, and with mammalian dipeptidases highly purified by others, shows remarkable similarities in molecular weights and molecular activities toward the preferred substrates but in the case of bacterial dipeptidase, differences in substrate specificities and in the effect of metal ions. Biochemistry, 1975 Sep 23, 14(19), 4221 - 6 S1 nuclease hydrolysis of single-stranded nucleic acids with partial double-stranded configuration; Rushizky GW et al.; The single-strand specific nuclease S1 from Aspergillus oryzae (EC 3.1.4.21) was purified 600-fold in 16% yield from dried mycelia . Determination of the isoelectric point of S1 nuclease as 4.3-4.4 allowed adjustment of chromatographic conditions such that the enzyme was isolated free of contaminating ribonucleases T1 and T2 . S1 nuclease so purified was used for removal of single-stranded portions from the RNA of the Escherichia coli phage MS2, which has a helical content of about 65% in vitro . At 23 degrees, increasing amounts of enzyme converted the RNA to mononucleotides in about equimolar base ratios . No small intermediates of chain length 2-8 were found . At 0 degrees, MS2 RNA hydrolysis was slower and reached, in exhaustive digests, a plateau where 70% of the substrate RNA remained insoluble in 66% EtOH . With {32P}MS2 RNA, strip chart counting of 6% acrylamide-6 M urea electrophoresis patterns of such digests gave recoveries of 80-91% in the form of defined oligomer bands . On 2.5% acrylamide-0.5% agarose gels, the molecular weights of the major oligomers were found to range from 25,000 to 41,000 . Similar to purified tRNAArg used as a control, these oligomers were not resistant to pancreatic RNase-RNase T1 hydrolysis at 37 degrees, and were not bound on hydroxylapatite at 50 degrees in 0.14 M sodium phosphate (pH 6.8) . Melting of the oligomers gave complex profiles without a clear Tm and showed an increase in A260 of 35% at 93 degrees over that at 28 degrees . Upon formaldehyde denaturation of MS2 RNA prior to S1 nuclease hydrolysis, no resistant oligomers were found. Biochemistry, 1975 Sep 23, 14(19), 4344 - 8 Molecular interactions between ribosomal proteins . Evidence for specificity of interaction between isolated proteins; Rohde MF et al.; The proteins S2, S3, S5, and S10 from the 30S ribosomal subunit of Escherichia coli was studied by analytical ultracentrifugation to characterize them in solution and to determine whether isolated protein-protein interactions exist . Such interactions, if specific, may therefore bear some relationship to the spatial organization of the subunit structure . It was found that protein S2 self-associates to a slight extent and that solution mixtures of S2 and S3 contain only enough dimeric species to account for the S2 dimer . Hence, no observable interaction was detected between S2 and S3 . Solution mixtures of proteins S5 and S10 revealed a species of molecular weight greater than either protein . The proposal is that S5 and S10 interact with an association equilibrium constant of 7.6 X 10(-5) M-1 at 3 degrees in a Tris buffer at pH 7.4 . It was also shown that solution with a 1:1:1 mixture by mass, of S2, S5, and S10 contained a species possessing a molecular weight consistent with a simple ternary complex of the three proteins. Biochemistry, 1975 Sep 23, 14(19), 4198 - 208 Structure of the modified nucleoside Q isolated from Escherichia coli transfer ribonucleic acid . 7-(4,5-cis-Dihydroxy-1-cyclopenten-3-ylaminomethyl)-7-deazaguanosine; Kasai H et al.; The structure of the unknown modified nucleoside Q, which is present in the first position of the anticodons of Escherichia coli tRNA Tyr, tRNA His, tRNA Asn, tRNA Asp, is proposed to be 7-(4,5-cis-dihydroxy-1-cyclopenten-3-ylaminomethyl)-7-deazaguanosine (1) . The structure of Q was deduced by means of its uv absorption, mass spectrometry, proton magnetic resonance spectroscopy, and studies of its chemical reactivity . The structure of Q is unique since it is a derivative of 7-deazaguanosine having cyclopentenediol in the side chain at the C-7 position . This is the first example of purine skeleton modification in a nucleoside from tRNA. Biochemistry, 1975 Sep 23, 14(19), 4185 - 91 Localization of the structural change induced in tRNA fMET (Escherichia coli) by acidic pH; Bina-Stein M et al.; We have compared the molecular mechanism of thermal unfolding for native tRNA fMet (Escherichia coli) and the denatured species produced by annealing at pH 4.3 . Relaxation kinetic measurements reveal that the transitions assigned to melting of TphiC, anticodon, and acceptor stem helices at neutral pH remain essentially unaltered at pH 4.3, but the transition corresponding to coupled melting of tertiary structure and dihydrouridine helix is greatly affected . The Tm of this region is more than 20 degrees higher at pH 4.3 and it has a larger enthalpy formation than in the native state . The transition dynamics are also considerably changed . In contrast to the native structure, tRNA fMet1 and tRNA fMet3 have similar tertiary structure stabilities at pH 4.3 . We conclude that the structural difference between native and acid-denatured forms is localized in the tertiary structure-dihydrouridine helix cooperative interaction region of the molecule. Biochim Biophys Acta, 1975 Sep 22, 403(1), 32 - 46 Circular dichroism studies of dihydrofolate reductase from a methotrexate-resistant strain of Escherichia coli B, MB 1428: ternary complexes; Greenfield NJ; Circular dichroism has been used to monitor the binding of pyridine nucleotide cofactors to enzyme-folate analog complexes of dihydrofolate reductase from Escherichia coli B (MB 1428) . The enzyme binds one molar equivalent of many folate analogs and two molar equivalents of several pyridine nucleotide cofactors . The apo-enzyme has very low optical activity . The binding of folate analogs including folate, dihydrofolate, methotrexate, trimethoprim and pyrimethamine induce large Cotton effects . Pyridine nucleotides when bound to the enzyme-folate analog complexes also induce new optically active bands; all the effects being due to the first molar equivalent of cofactor bound . NADPH and NADP+ induce very similar bands when bound to the enzyme-methotrexate complex suggesting that the geometry of the complexes formed are very similar . The oxidized and reduced cofactor likewise have similar effects on the enzyme-folate complex . However, NADPH and NADP+ addition to both the enzyme-trimethoprim and enzyme-pyrimethamine complexes have significantly different effects on the circular dichroism spectra, suggesting that the inhibitors which are less homologous to the natural dihydrofolate substrate allow more conformational freedom in the enzyme-inhibitor-cofactor complex . In most cases the prior binding of the folate analog greatly increases the binding of the first molar equivalent of cofactor so that at concentrations of approx . 5-20 muM the binding appears stoichiometric . Pyrimethamine is an exception in that it apparently has no effect on the binding of NADPH to the enzyme. Biochim Biophys Acta, 1975 Sep 22, 403(1), 221 - 31 Studies on aspartase . II . Role of sulfhydryl groups in aspartase from Escherichia coli; Mizuta K et al.; Aspartase (L-aspartate ammonia-lyase, EC 4.3.1.1) of Escherichia coli W contains 38 half-cystine residues per tetrameric enzyme molecule . Two sulfhydryl groups were modified with N-ethylmaleimide or 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) per subunit, while 8.3 sulfhydryl groups were titrated with p-mercuribenzoic acid . In the presence of 4 M guanidine - HCl, 8.6 sulfhydryl groups reacted with DTNB per subunit . Aspartase was inactivated by various sulfhydryl reagents following pseudo-first-order kinetics . Upon modification of one sulfhydryl group per subunit with N-Ethylmaleimide, 85% of the original activity was lost; a complete inactivation was attained concomitant with the modification of two sulfhydryl groups . These results indicate that one or two sulfhydryl groups are essential for enzyme activity . L-Aspartate and DL-erythro-beta-hydroxyaspartate markedly protected the enzyme against N-ethylmaleimide-inactivation . Only the compounds having an amino group at the alpha-position exhibited protection, indicating that the amino group of the substrate contributes to the protection of sulfhydryl groups of the enzyme . Examination of enzymatic properties after N-ethylmaleimide modification revealed that 5-fold increase in the Km value for L-aspartate and a shift of the optimum pH for the activity towards acidic pH were brought about by the modification, while neither dissociation into subunits nor aggregation occurred . These results indicate that the influence of the sulfhydryl group modification is restricted to the active site or its vicinity of the enzyme. Biochim Biophys Acta, 1975 Sep 22, 403(1), 208 - 20 Studies of homogeneous "biosynthetic" L-threonine dehydratase from Escherichia coli K-12 . Some kinetic properties and molecular multiplicity; Kagan ZS et al.; "Biosynthetic" L-threonine dehydratase (EC 4.2.1.16) was purified to a homogeneous state with 29% yield of total activity from Escherichia coli K-12 . The homogeneity of the enzyme was shown by polyacrylamide gel disc electrophoresis in the presence of dodecyl sulphate . The enzyme consisted of equal subunits having a molecular weight of about 57 000 . The polyacrylamide gel disc electrophoresis has shown that the native enzyme consisted of a set of oligomeric forms . The multiplicity of molecular organization of the enzyme was reflected in complicated kinetic behaviour: at pH greater than 9 on the plots of initial reaction rate (v) versus initial substrate concentration ({S}o) there were four inflexion points (two intermediate plateaux), the position and deepness of which depended on enzyme concentration . At pH 8.3 on the v versus {S}o plots appeared two inflexion points (one intermediate plateu), the position of which practically did not depend on enzyme concentration in the reaction mixture, but strongly depended on the enzyme concentration in the stock solution . Repeated polyacrylamide gel disc electrophoresis of several oligomeric forms, isolated by the first electrophoresis, has shown that the oligomeric forms underwent a slow polymerization . It was suggested that "biosynthetic" L-threonine dehydratase from E . coli K-12 is a set of multiple oligomeric forms, having different kinetic parameters . Probably, each form of the enzyme has a "simple" kinetics characterized by hyperbolic or sigmoidal shape of v versus {S}o plots . The rate of equilibrium installation between the oligomeric forms was small in comparison with the enzyme reaction velocity, that lead to the complex kinetic curves, appearing as a result of summing up of the kinetics inherent to theindividual forms. J Biol Chem, 1975 Sep 10, 250(17), 7074 - 7 Thermal regulation of the membrane lipid composition of Escherichia coli . Evidence for the direct control of fatty acid synthesis; Cronan JE Jr; Starvation of strains of Escherichia coli which are glycerol auxotrophs and are also defective in beta oxidation results in the accumulation of large amounts of free fatty acid (Cronan, J . E., Jr., Weisberg, L . W., and Allen, R . G . (1975) J . Biol . Chem . 250, 5835-5840) . We now report that the ratio of saturated to unsaturated species appearing in the free fatty acid fraction depends on the incubation temperature at the time of synthesis of these acids . This result indicates that fatty acid synthesis is one site of the thermal control of phospholipid fatty acid composition . We also report experiments on the incorporation of exogenously supplied fatty acids into membrane phospholipids . The effect of temperature on this incorporation supports the hypothesis that a second site of thermal regulation acts at the level of phosphatidic acid synthesis. Nature, 1975 Sep 18, 257(5523), 193 - 7 In vitro synthesis of tRNA precursors and their conversion to mature size tRNA; Daniel V et al.; Two Escherichia coli tRNA gene clusters, tRNA1Tyr (su3+ and su3-) and tRNA2Tyr, tRNA2Gly (su+36), tRNA3Thr, were transcribed in a purified in vitro system . Evidence indicates that the adjacent tRNA genes are transcribed together as a common precursor of large size, which, on incubation with crude cell extracts, yields mature tRNA molecules. Biochim Biophys Acta, 1975 Sep 16, 406(1), 50 - 9 Inducible gluconate permease in a gluconate kinase-deficient mutant of Escherichia coli; Robin A et al.; Gluconate-resistant mutants were isolated from Escherichia coli strain DF 1070 deficient in phosphogluconate dehydrogenase (EC 1.1.1.44) and in phosphogluconate dehydrogenase (EC 4.2.1.12) which is inhibited by gluconate . Among the resistant mutants, AR 13 has been identified as a gluconate kinase (EC 2.7.1.12)-deficient strain . This mutant exhibits an inducible gluconate transport system capable of concentrating gluconate in the cytoplasm against a concentration gradient . The accumulated gluconate is subject to permanent turnover, and is not chemically modified . The kinetics of induction and deinduction indicate a single inducible component, rate limiting for the transport function, and the distribution of transport capacity among non-induced progeny of induced parents indicates that the inducible protein is membrane bound. Biochim Biophys Acta, 1975 Sep 16, 406(1), 36 - 49 A novel electrophoretic fractionation of Escherichia coli envelopes; Joseleau-Petit D et al.; Particulate fractions of Escherichia coli have been submitted to electrophoretic fractionation in a buffer stabilized by sucrose gradient . Inner membrane and outer membrane were readily resolved . A combination of electrophoresis, fractional centrifugation and gel filtration can remove remaining contamination by ribosomes and cytoplasm . The presence of particles containing no phospholipids was detected after differential centrifugation . The nature of this fraction is unknown . The inner membrane exhibited heterogeneity on electrophoresis. Mol Gen Genet, 1975 Sep 15, 140(1), 81 - 90 Catabolite repression in Escherichia coli K12 mutants defective in glucose transport; Gershanovitch VN et al.; The phenomenon of glucose catabolite repression was studied in Escherichia coli mutants unable to transport this carbohydrate . Th