J Biol Chem, 1976 Jul 25, 251(14), 4208 - 13 Identification of sn-glycero-1-phosphate and phosphoethanolamine residues linked to the membrane-derived Oligosaccharides of Escherichia coli; Kennedy EP et al.; A previous report from this laboratory (van Golde, L.M.G., Schulman, H., and Kennedy, E.P . (1973) Proc . Natl . Acad . Sci . U.S.A . 70, 1368-1372) described the discovery in Escherichia coli of a novel class of oligosaccharides, containing glucose as the sole sugar, substituted with glycerophosphate units derived from membrane phospholipids, and with succinic acid in O-ester linkage . These membrane-derived oligosaccharides, comprising about 0.5 to 1.0% of the dry weight of E . coli, represent a family of closely related oligosaccharides that may be subfractionated on anion exchange resins . The present paper describes studies of the oligosaccharide A-1 described by van Golde et al . in the previous report . The glycerophosphate linked to the oligosaccharide in phosphodiester bond is the sn-glycero-1-P enantiomer . This finding strongly supports the previous conclusion that the oligosaccharides are the acceptors of the polar headgroups of membrane phospholipids, since the unesterified glycerophosphate of phosphatidyl glycerol is an sn-glycero-1-P residue, otherwise rare in nature . The glycerophosphate residues in the membrane-derived oligosaccharide are not substituted in the sn-2 or sn-3 positions, since they are readily oxidized by periodate under mild conditions . Alkaline hydrolysis liberates glycerophosphate, and only negligible amounts of free glycerol, consistent with the view that the glycerophosphate residues are linked to glucose units through position 6, unfavorable for the formation of glucose cyclic phosphate intermediates that would eliminate free glycerol . Oligosaccharide A-1 (but not Fraction A-2) contains phosphoethanolamine residues equivalent to 30 to 40% of the total phosphorus . The phosphoethanolamine residues are linked to position 6 of glucose units, as proved by the isolation of glucose 6-phosphate as a product of partial acid hydrolysis. Lancet, 1976 Jul 24, 2(7978), 188 - 92 Host resistance to lipopolysaccharides in the pathogenesis of multiple sclerosis and membranoproliferative glomerulonephritis; Back U et al.; Host resistance against bacterial lipopolysaccharides (L.P.S.) and especially against its toxic part lipid A has earlier been demonstrated in biological assays . In this paper an aryl-esterase is shown to be associated with alfa-1-lipoprotein (ArE) and is probably responsible for the detoxification of L.P.S . in man . Furthermore C3 is shown to be activitated by L.P.S . From these facts it is suggested that ArE performs the initial degradation of L.P.S . followed by complement activation and trapping of the L.P.S.--complement complex in the reticuloendothelial system . It is postulated that a deficient host response against L.P.S . can be the triggering mechanism in multiple sclerosis due to the lack of ArE in myelin, and that an infectious-agent/L.P.S . syndrome can activate latent infections in connection with a severe hyperreactivity to L.P.S . Preliminary investigations in patients with membranoproliferative glomerulonephritis have shown low levels of ArE in serum . This change, together with the low C3 values in these patients, may result in deficient L.P.S . detoxification and it is suggested that L.P.S . are at least partly responsible for the production of C3 nephritic factor. Mol Gen Genet, 1976 Jul 23, 146(2), 107 - 15 Genetic exchanges caused by ultraviolet photoproducts in phage lambda DNA molecules: the role of DNA replication; Lin PF et al.; Genetic recombination induced by structural damage in DNA molecules was investigated in E . coli K12 (lambda) lysogens infected with genetically marked phage lambda . Photoproducts were induced in the phage DNA before infection by exposing them either to 313 nm light in the presence of acetophenone or to 254 nm light . To test the role of the replication of the damaged phage DNA on the frequency of the induced recombination, both heteroimmune and homimmune crosses were performed . First, samples of a heteroimmune phage lambda imm434 P80 exposed to these treatments were allowed to infect cells lysogenic for prophage lambda cI857 P3 . Phage DNA replication and maturation took place, and the resulting progeny phages were assayed for the frequency of P+ recombinants . Recombination was less frequent in infected cells exposed to visible light and in wild type cells able to perform excision repair than in excision-defective lysogens . Therefore, much of the induced recombination can be attributed to the pyrimidine dimers in the phage DNA, the only photoproducts known to be dissociated by photoreactivating enzyme . Second, in homoimmune crosses, samples of similarly treated homoimmune lambda P3 phages were allowed to infect lysogens carrying lambda cI857 P80 . Replication of the phage DNA containing ultraviolet photoproducts was repressed by lambda immunity, and was further blocked by the lack of the P gene product needed for replication . The lysogens were purified and scored for both colony forming ability and for P+ recombinant prophages . The 254 nm photoproducts increased the frequency of recombination in these homimmune crosses, even though phage DNA replication was blocked . Irradiation with 313 nm light and acetophenone M, which produces dimers and unknown photoproducts, was not as effective per dimer as the 254 nm light . It is concluded from these results that certain unidentified 254 nm photoproducts can cause recombination even in the absence of DNA replication . They are not pyrimidine dimers, as they are not susceptible to excision repair or photoreactivation . In contrast, pyrimidine dimers appear to cause recombination only when the DNA containing them undergoes replication. Mol Gen Genet, 1976 Jul 23, 146(2), 139 - 45 Multiple regulation of nucleoside catabolizing enzymes: effects of a polar dra mutation on the deo enzymes; Albrechtsen H et al.; Strains with an amber, polar mutation in the dra1 gene have been isolated . The mutation was introduced into a set of isogenic strains, wild type or with concurrent regulatory mutations, and further characterized by suppression and heat inactivation experiments . The effect of the polar dra mutation on the three remaining genes of the deo operon, the tpp, drm and pup genes, was determined by estimating the enzyme levels in the various dra-mutants . The effect was found to be non-coordinate, indicating the formation in the cells of two types of transcripts: A tetracistronic unit, containing the message from all four genes, and a dicistronic unit, covering the two distal genes only. Mol Gen Genet, 1976 Jul 23, 146(2), 199 - 207 The construction in vitro of transducing derivatives of phage lambda; Borck K et al.; Methods are described for the construction of plaque-forming, transducing derivatives of phage lambda, using appropirate receptor genomes and fragments of DNA generated by the restriction enzymes endo R.EcoRI and endo R.HindIII . The general properties of the transducing derivatives are described and discussed . Plaque-forming phages carrying the E . coli trp, his, cysB, thyA, supD, supE, sup F, hsd, tna and lig genes have been isolated. Mol Gen Genet, 1976 Jul 23, 146(2), 179 - 88 Initial trp operon sequence in Escherichia coli is transcribed without coupling to translation; Kano Y et al.; The transcription of the "leader" region (Bronson et al., 1973) of the trp operon in Escherichia coli was studied in normal mutants which delete most of the operator-distal region of the operon {a deletion strain (trp OAEG) retaining only about one third of the "leader" region and two deletion strains (trpOAE14 and trpOAE2) retaining the whole "leader" region and an initial portion of the trpE}, as well as in a strain with an intact trp operon, but with a temperature-sensitive lesion in ribosomal protein factor EFTs (strain HAK88) . In these deletion mutants, mRNA molecules corresponding to the "leader" region were detected as most of the trp-specific mRNA . Less inhibition of transcription, of the promoter-proximal portion of the trp "leader" region than that of more distal genes of the operon, was found in chloramphenicol-treated cells of strain trpOAE14 . It was also observed that transcription of the initial one third portion of the "leader" region was not repressed by tryptophan in strains trpOAE6 and trp OAE14 . A similar effect of a translation block on transcription of the distal part of the "leader" region was observed with strain HAK88 at the nonpermissive temperature . In sedimentation analysis of polyribosomes containing the trp mRNA molecules from the deletion mutants, trp mRNA from strain trpOAE14 was found in monosomes and small polyribosomes, whereas the majority of the trp mRNA from strain trpOAE6 was found joined to a single ribosome or ribosomal subunit . These results suggest that ribosomes bind in vivo to a site(s) located in the middle of the "leader" mRNA sequence, and that the initial transcription of the trp operon does not require any connection to functional translational machinery, while continuation of RNA synthesis beyond a first ribosome binding site seems indispensably coupled to ribosome function. Mol Gen Genet, 1976 Jul 23, 146(2), 161 - 5 Effect of tra mutations on F factor-specified immunity to lethal zygosis; Skurray RA et al.; Hfr, F+, and F-prime cells are, unlike F- cells, insensitive to an excess of Hfr donor cells, indicating that there is an F factor mediated immunity to lethal zygosis (I1z) . Results with Flac episomes carrying traJ, traS or various polar mutations in the tra region indicate that this immunity is independent of surface exclusion, of traJ control, and of all known genes within the tra operon . However, analysis of a series of strains with deletions in the F factor, extending from the right into the tra region, suggests that a gene for immunity to lethal zygosis is located within the tra region . We therefore conclude that I1z is genetically complex, and present a hypothesis to account for these results. Biochim Biophys Acta, 1976 Jul 20, 441(1), 38 - 47 Distribution of lipids in cytoplasmic and outer membranes of Escherichia coli K12; Lugtenberg EJ et al.; The lipid composition of cytoplasmic and outer membranes of Escherichia coli K12 was studied . Compared with the cytoplasmic membrane, the outer membrane is enriched in both saturated fatty acids and phosphatidylethanolamine . This is also the case when the fatty acid composition of the phospholipids is changed, either by varying the growth temperature or by using mutants with alterations in their fatty acid metabolism . Phosphatidylethanolamine of the outer membrane contains relatively more saturated fatty acids than phosphatidylethanolamine of the cytoplasmic membrane. Biochim Biophys Acta, 1976 Jul 16, 435(4), 349 - 61 Synthesis of inducible enzyme in Escherichia coli recovering from prolonged energy starvation; Hwang LT et al.; A marked breakdown of ribosomes and rRNA occurs in Escherichia coli cells during prolonged deprivation of a carbon source (energy starvation) . In E . coli recovering from energy starvation: (a) synthesis of RNA started immediately, total protein synthesis showed a delay of 5 to 10 minutes; (b) beta-galactosidase, tryptophanase and serine deaminase could not be induced in the first 50--70 min; (c) a lag of 60 min in the synthesis of beta-galactosidase was observed in a lac constitutive mutant of E . coli; synthesis of the constitutive enzyme malate dehydrogenase did not shown any delay . RNA synthesized in the early stages of recovery contained a higher percentage of low molecular weight molecules than RNA synthesized after 70 min of recovery or during exponential growth . Messenger RNA specific for beta-galactosidase was not synthesized for the first 50--60 min of recovery even when the specific inducer was added to the cultures. Eur J Biochem, 1976 Jul 15, 66(3), 583 - 9 1 . Membrane vesicles of Escherichia coli K-12 CS7, a strain gentically derepressed for glutamate permease, maintain low aspartate transport activity, like that of prep; Kahane S et al.; 1 . Membrane vesicles of Escherichia coli K-12 CS7, a strain gentically derepressed for glutamate permease, maintain low aspartate transport activity, like that of preparations of the wild-type parent . Growth of the parent CS101 on aspartate as the source of carnon or nitrogen results in derepression of both asparatate and glytamate transport . Growth of strain CS7 on aspartate derepresses aspartate transport to the same extent as in strains CS101, but only slightly increases the derepressed level of glutamate transport activity . 2 . The affinity of the membrane transport system for glutamate is enhanced by sodium, while that for asparate is not . 3 . Although the affinities for glutamate (23 muM) and aspartate (12 muM) are similar, aspartate does not inhibit glutamate transport, while glutamate competitively inhibits aspartate transport . 4 . Aspartate transport, but not glutamate transport, is competitively inhibited by C4 dicarboxylic acids, whereas 2-oxoglutarate competitively inhibits glutamate transport, but not aspartate transport . 5 . Competitive inhibition of L-aspartate transport by L-glutamate and by the 5-methyl ester of L-glutamate is abolished in the presence of 2-oxoglutarate . However, 2-oxoglutarate does not affect the competitive inhibition of L-aspartate transport by D-aspartate and by DL-threo-3-hydroxyaspartate . The relationship between the two dicarboxylic amino acid transport systems and the spatial characteristics of the aspartate carrier are discussed in the light of these findings. Biochim Biophys Acta, 1976 Jul 15, 436(4), 800 - 10 Membrane-associated reactions in ubiquinone biosynthesis in Escherichia coli . 3-Octaprenyl-4-hydroxybenzoate carboxy-lyase; Leppik RA et al.; A sensitive and quantitative assay for 3-octaprenyl-4-hydroxybenzoate carboxy-lyase has been developed . This enzyme, which catalyses the third reaction in ubiquinone biosynthesis in Escherichia coli, was partially purified and some of its properties determined . It was found that a considerable proportion of the carboxylyase activity could be separated from the membrane fraction in cell extracts prepared using a French press . Gel filtration showed the molecular weight of the enzyme to be about 340 000 . For optimal activity the carboxy-lase was shown to require Mn2+, washed membranes or an extract of phospholipids, and an unidentified heat stable factor of molecular weight less than 10 000 . The carboxy-lyase reaction was also shown to be strongly stimulated by dithiothreitol and methanol . The properties of the carboxy-lyase are compared with the three other enzymes concerned with ubiquinone biosynthesis in E . coli which have been studied in vitro . The fact that the substrate of the carboxy-lyase is membrane-bound and the enzyme is stimulated by phospholipid suggests that it normally functions in association with the cytoplasmic membrane in vivo. Eur J Biochem, 1976 Jul 15, 66(3), 443 - 6 Dinitrophenol, dicoumarol and pentachlorophenol as inhibitors and parasite substrates in the ATP phosphoribosyltransferase reaction; Dall-Larsen T et al.; Adenosine-triphosphate phosphoribosyltransferase from Escherichia coli is inhibited by dicoumarol and pentachlorophenol in competition with ATP . Ki was approximately 60 muM for dicoumarol and 50 muM for pentachlorophenol . Carbonylcyanide m-chlorphenylhydrazine did not seem to have any kinetic effect . Dicoumarol is bound to the extent of 6 sites per enzyme hexamer with a dissociation constant Kd of 50 muM . Dicoumarol and pentachlorophenol partly prevent the binding of ATP and AMP to the transferase . The reverse reaction is inhibited by dicoumarol and pentachlorophenol without changes in {s}0.5 for phosphoribostladenosine trophosphate . Dicumarol, dinitrophenol and pentachlorophenol diminish the yield of phosphoribosyladenosine triphosphate in the transferase reaction apparently by acting as parasite substrates; carbonylcyanide m-chlorophenylhydrazone had no effect. Biochemistry, 1976 Jul 13, 15(14), 3031 - 9 Regions of tRNA important for binding to the ribosomal A and P sites; Sprinzl M et al.; Studies on the enzymatic inhibition of phenylalanyl-tRNAPhe and formylmethionyl-tRNAFMet binding to the ribosomes by defined tRNA fragments indicate that beside the anticodon the following regions of tRNA are important for ribosomal A-site interaction: the TppsipCp sequence, the CpCpA end, and hU loop . In contrast, binding to the ribosomal P site is not inhibited by the fragments of uncharged yeast tRNAPhe containing the hU or the TpsiC loop of the molecule . Comparative studies on the inhibitory effect of the oligonucleotides TppsipCpGp and UpUpCpGp indicate that the presence of the minor bases in TpsiC loop is not an essential prerequisite for the binding of tRNA to the ribosomal A site . Furthermore, evidence is presented that shows that the binding of the TppsipCpGp oligonucleotide to the ribosomes influences the ribosomal P site and increases there the efficiency of the codon-anticodon interaction . It is suggested that the TppsipCpGp binds to the ribosomal A site and competes there with the TpsiC loop of the aminoacyl-tRNA for the same binding site . A model for the interaction between tRNA and the ribosomal A site is proposed that involves partial unfolding of hU and TpsiC loops of the tRNA and, therefore, suggests the dynamic involvement of tRNA in protein synthesis. Biochemistry, 1976 Jul 13, 15(14), 2986 - 94 Modification of membrane lipid: physical properties in relation to fatty acid structure; Baldassare JJ et al.; Differential scanning calorimetry (DSC) and electron spin resonance (ESR) measurements were made to characterize how modifications in the fatty acid composition of Escherichia coli affected the thermotropic phase transition(s) of the membrane lipd . When the fatty acid composition contained between 20 and 60% saturated fatty acids, the DSC curves for isolated phospholipids and cytoplasmic membranes showed a broad (15-25 degree C) gel to liquid-crystalline phase transition, the position of which depended on the particular fatty acid composition . Utilizing multiple lipid mutants, enrichment of the membrane phospholipids with a single long-chain cis-monoenoic fatty acid in excess of that possible in a fatty acid levels less than 20% and gradually replaced the broad peak as the cis-monoenoic fatty acid content increased . These results were obtained with phospholipids, cytoplasmic membranes, and whole cells . With these same phopholipids, plots of 2,2,6,6-tetramethylpiperidinyl-1-oxy partitioning and ESR order parameters vs . 1/T revealed discontinuities at temperatures 40-60 degrees C above the calorimetrica-ly measured gel to liquid-crystalline phase transitions . Moreover, when the membrane phospholipids were enriched with certain combinations of cis-monenoic fatty acids (e.g., cis-delta 9-16:1 plus cis-delta 11-18:1) the DSC curve showed a broad gel to liquid crystalline phase change below 0 degrees C but the ESR studies revealed no discontinuities at temperatures above those of the gel to liquid-crystalline transition . These results demonstrated that enrichment of the membrane lipids with molecules in which both fatty acyl chains are identical cis-monoenoic residues led to a distinct type of liquid-crystalline phase . Furthermore, a general conclusion from this study is that Escherichia coli normally maintains a heterogeneous mixture of lipid molecules and, by so doing, prevents strong lipid-lipid associations that lead to the formation of lipid domains in the membrane. J Biol Chem, 1976 Jul 10, 251(13), 4090 - 4 On the processive mechanism of Escherichia coli DNA polymerase I . Delayed initiation of polymerization; Bambara RA et al.; Escherichia coli DNA polymerase I shows a delay in the initiation of polymerization after binding to the cohesive ends of lambda DNA . This delay is significantly longer than the time required for the synthesis of an octanucleotide sequence on the right-hand cohesive end of the lambda DNA . When the extent of polymerization is limited by omission of one or more of the deoxynucleoside triphosphates, and polymerization started again by their addition, the delay still occurs . A plausible explanation for this phenomenon is that two forms of the enzyme or enzyme - DNA complex exist, only one of which is active . The delay, therefore, represents the time necessary to convert the inactive to the active form of the enzyme or enzyme complex . One consequence of the defect in DNA polymerase I, due to the polA12 mutation, is apparently to alter the equilibrium between the two forms . However, the rate of polymerization and the rate of conversion of inactive to active enzyme or enzyme complex are not changed significantly. J Biol Chem, 1976 Jul 10, 251(13), 4085 - 9 Biochemical characterization of mutant forms of DNA polymerase I from Escherichia coli . II . The polAex1 mutation; Uyemura D et al.; DNA polymerase I has been purified to homogeneity from an Escherichia coli K12 strain bearing the temperature-sensitive conditionally lethal mutation, polAex1 . The purified enzyme shows no defect in its polymerase or 3' leads to 5'-exonuclease activities; however, its 5' leads to 3'-exonuclease activity is abnormally low at both 30 degrees and 43 degrees . Although the mutant enzyme is able to catalyze the coordinated 5' leads to 3' polymerization and 5' leads to 3' exonucleolytic hydrolysis of nucleotides at a nick in duplex DNA ("nick translation") at a measurable rate at 30 degrees, this reaction is undetectable at 43 degrees . This defect is very likely responsible for the retarded joining of nascent DNA fragments and the consequent loss of viability that occur in the mutant at this temperature. J Biol Chem, 1976 Jul 10, 251(13), 4078 - 84 Biochemical characterization of mutant forms of DNA polymerase I from Escherichia coli . I . The polA12 mutation; Uyemura D et al.; DNA polymerase I has been purified to greater than 90% homogeneity from a strain of Escherichia coli K12 that bears the temperature-sensitive DNA polymerase I mutatation, polA12 . The mutant enzyme has a reduced electrophoretic mobility and sedimentation rate . It is abnormally thermolabile and is rapidly inactivated at low salt concentrations . Its polymerase and 5' leads to 3' exonuclease activities are not grossly defective at 30 degrees, yet its capacity to promote the concerted 5' leads to 3' polymerization and the 5' leads to 3' exonucleolytic hydrolysis of nucleotides at a nick ("nick translation") is decreased 10-fold . These effects are probably the result of a significant alteration in the tertiary structure of the enzyme. J Biol Chem, 1976 Jul 10, 251(13), 4071 - 7 Sea urchin sperm guanylate cyclase . Purification and loss of cooperativity; Garbers DL; The Lubrol-dispersed guanylate cyclase from sea urchin sperm was purified and isolated essentially free of detergent by GTP affinity chromatography, DEAE-Sephadex chromatography, and gel filtration . After removal of the detergent, the enzyme remained in solution in the presence of 20% glycerol . The specific activity of the purified enzyme was about 12 mumol of guanosine 3':5'-monophosphate (cyclic GMP) formed - min-1 - mg of protein-1 at 30 degrees, an activity about 4600 times that of a soluble guanylate cyclase purified recently from Escherichia coli (Macchia V., Varrone, S., Weissbach, H., Miller, D.L., and Pastan, I . (1975) J . Biol . Chem . 250, 6214-6217) . The cyclic GMP phosphodiesterase activity was negligible and adenosine 3':5'-monophosphate (cyclic AMP) phosphodiesterase was not detectable in the purified preparation . Cyclic AMP formation from ATP occurred at a rate of 0.002% of that of guanylate cyclase . In the absence of phosphodiesterase or guanosine triphosphatase inhibitors, 100% of the added GTP was converted to cyclic GMP . The purified enzyme required Mn2+ for maximum activity, the relative rates in the presence of Mg2+ or Ca2+ being less than 0.6% of the rates with Mn2+ . The purified enzyme displayed classical Michaelis-Menten kinetics with respect to MnGTP (apparent Km is approximately equal to 170 muM) in contrast to the positively cooperative kinetic behavior displayed by the unpurified, detergent-dispersed, or particulate guanylate cyclase . The molecular weight of the purified enzyme was approximately 182,000 as estimated on Bio-Gel A-0.5m columns equilibrated in the presence or absence of 0.1 M NaCl . The unpurified, detergent-dispersed enzyme also migrated with an apparent molecular weight of 182,000 on columns equilibrated with 0.5% Lubrol WX and 0.1 M NaCl, but it migrated as a large aggregate (molecular weight is greater than 5 X 10(5)) on columns equilibrated in the absence of either the detergent of NaCl . After gel filtration, the unpurified, dispersed enzyme still yielded positive cooperative kinetic patterns as a function of MnGTP . Na dodecyl-SO4 gel electrophoresis of the enzyme after the DEAE-Sephadex or the gel filtration steps resulted in two major protein bands with estimated molecular weights of 118,000 and 75,000 . Whether or not these protein bands represent the subunit molecular weights of guanylate cyclase is unknown at present. Biochim Biophys Acta, 1976 Jul 8, 438(2), 563 - 73 Purification and properties of tyrosine-sensitive 3-deoxy-D-arabino-heptolosonate-7-phosphate synthetase of Escherichia coli K12; Dusha I et al.; 1 . The tyrosine-sensitive allosteric first enzyme of the aromatic amino acid biosynthetic pathway, 3-deoxy-D-arabinoheptulosonate 7-phosphate synthetase (7-Phospho-2-keto-3-deoxy-D-arabino-heptonate D-erythrose 4-phosphate-lyase (pyruvate phosphorylating), EC 4.1.2.15) has been purified from a mutant strain of Escherichia coli . 2 . The enzyme activity was inhibited to 50% at 2-10(-5) M tyrosine and to 90% at 2-10(-4) M tyrosine concentration . At tyrosine concentrations lower than 2-10(-5) M a cooperative interaction between tyrosine binding sites was observed . 3 . Co2+ increased the enzyme activity about 2-2.5-fold . The presence of Co2+ ions stabilized the enzyme . EDTA inhibited the enzyme activity, and this inhibition was reversed by Co2+ . Tyrosine-sensitive DAHP synthetase seems to be a metal containing enzyme . 4 . Kinetic experiments were carried out to study the catalytic action . Contrary to earlier suggestions it is concluded, that the reaction mechanism appears to be more complex--with either the ping-pong or sequential type predominating, depending on conditions. Biochim Biophys Acta, 1976 Jul 8, 438(2), 487 - 94 Kinetic properties and the effect of substrate analogues on 5'-methylthioadenosine nucleosidase from Escherichia coli; Ferro AJ et al.; 5'-Methylthioadenosine nucleosidase (EC 3.2.2-) from Escherichia coli has been purified 220-fold . A molecular weight of 31 000 for the enzyme was estimated from gel filtration on Sephadex G-150 . The Km for 5'-methylthioadenosine was 3.1-10(-7) M . In addition to 5'-methylthioadenosine, the nucleoside analogues 5'-ethylthioadenosine, 5'-n-propylthioadenosine, and S-adenosyl-homocysteine also served as substrates for the enzyme . These substrate analogues acted as competitive inhibitors of the reaction with 5'-methylthioadenosine . The Ki values for 5'-ethylthioadenosine, 5'-n-propylthioadenosine, and S-adenosylhomocysteine were determined to be 1.3-10(-7) M, 4.6-10(-8) M, and 1.92-10(-7) M respectively. Mol Gen Genet, 1976 Jul 5, 146(1), 79 - 83 Rifampicin-induced protein synthesis: A pre-requisite for increased expression of the beta beta' operon in Escherichia coli; Tittawella PB; In a rifS/rifR heterodiploid strain of E . coli, a 4 minute pulse of rifampicin can induce a prolonged (greater than 60 min) increase in the rate of synthesis of the RNA polymerase subunits, beta and beta' . The application of a constraint on the fidelity of protein synthesis during, but not after, the rifampicin pulse partially arrests the development of this capacity for subunit synthesis . I discuss the implications of these findings in relation to the control of the BB' operon in E . coli. Biochim Biophys Acta, 1976 Jul 2, 435(3), 306 - 14 An in vivo effect of the metabolites L-alanine and glycyl-L-leucine on the properties of the lysyl-tRNA synthetase from Escherichia coli K-12 . II . Kinetic evidence; Hirshfield IN et al.; Wild-type Escherichia coli K-12 was grown in minimal medium alone or with the addition of 20 mM L-alanine or 3 mM glycyl-L-leucine . A lysyl-tRNA synthetase mutant strain was grown in minimal medium containing 20mM L-alanine . The lysyl-tRNA synthetase from these strains was purified to 70-90% of homogeneity . Kinetic studies comparing the effect of thermal and urea inactivation on these different lysyl-tRNA synthetase preparations and measurement of the Michaelis constant for lysine and transfer RNA indicated that growth of Escherichia coli in the presence of alanine and glycyl-L-leucine induces an alteration in the properties of the synthetase . Measurement of the apparent Km for ATP at pH 7.25 indicates lysyl-tRNA synthetase has two two binding sites for this substrate, and further studies indicated a dependence of the apparent Km for lysine on the ATP concentration. Biochim Biophys Acta, 1976 Jul 2, 435(3), 290 - 305 An in vivo effect of the metabolites L-alanine and glycyl-L-leucine on the properties of lysyl-tRNA synthetase from Escherichia coli K-12 . I . Influence on subunit composition and molecular weight distribution; Hirshfield IN et al.; Lysyl-tRNA synthetase was purified to 70-90% of homogeneity from Escherichia coli K-12 . The enzyme was purified from wild-type cells grown in minimal medium, or minimal medium containing either 20 mM L-alanine or 3 mM glycly-L-leucine . The synthetase was similarly purified from a mutant strain grown in minimal medium plus 20 mM L-alanine . Results based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, gel filtration, and trypsin inactivation studies indicate (A) that the presence of L-alanine of glycyl-L-leucine in the culture medium alters the properties of the wild-type enzyme; (B) that the alteration of the synthetase by l-alanine and glycyl-L-leucine is different; and (c) that the molecular weight of lysyl-tRNA synthetase is at least 135000--140000 . The results suggest that most likely the metabolites modify the structure of lysyl-tRNA synthetase, but the possibility that the metabolites induce the synthesis of a new lysyl-tRNA synthetase cannot be completely eliminated. Biochim Biophys Acta, 1976 Jul 2, 435(3), 251 - 7 Undermethylated transfer RNA does not support phage RNA-directed in vitro protein synthesis; Stulberg MP et al.; A cell-free protein synthesis system from Escherichia coli Q13 was depleted of mRNA and tRNA so that restoration of maximum activity was dependent of the addition of these components . Protein synthesis, directed by either MS2 or Qbeta phage RNA, was stimulated significantly by the addition of normal tRNA from either E . coli Q13 or B . In contrast, undermethylated tRNA from methionine-starved E . coli RCrel did not cause this stimulation . It is concluded that undermethylated tRNA Lacks sufficient base modifications to function in protein synthesis. Science, 1976 Jul 2, 193(4247), 60 - 2 Enhanced template activity in chromatin from adrenal medulla after phosphorylation of chromosomal proteins; Chuang DM et al.; Translocation of protein kinase to the nucleus had been implicated earlier in the transsynaptic control of gene expression mediated by cholinergic nerves in adrenal medulla . Phosphorylation of chromosomal proteins by adenosine 3',5'-monophosphate-dependent protein kinase and adenosine 3',5'-monophosphate enhances the template activity of chromatin from adrenal medulla . When homologous RNA polymerase II is used the relative activation is greater than that obtained with Escherichia coli RNA polymerase . The substrate for such phosphorylation does not seem to be RNA polymerase II . Phosphorylation of specific acidic protein probably mediates this enhancement of template activity. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2351 - 5 Nucleotide sequence of region preceding trp mRNA initiation site and its role in promoter and operator function; Bennett GN et al.; The nucleotide sequence of the region preceding the transcription initiation site of the tryptophan operon of Escherichia coli was determined . Essentially all of the trp operator precedes the transcribed portion of the operon . The deduced sequence contains the recognition site of endonuclease Hpa I . This site is protected from Hpa I cleavage by RNA polymerase and by trp repressor . Regions of 2-fold symmetry are present in the DNA sequence. Lab Anim, 1976 Jul, 10(3), 199 - 202 Histological findings in rabbits which died with symptoms of mucoid enteritis; Meshorer A; Histological and aetiological similarities between rabbit mucoid enteritis and human ulcerative colitis are briefly discussed . Escherichia coli seems to be associated with the rabbit disease, and treatment aimed at this organism has been followed by a period of 4 years free from mucoid enteritis. Mol Biol (Mosk), 1976 Jul-Aug, 10(4), 715 - 24 Structure of animal mitochondrial DNA: nucleotide composition, pyrimidine clusters, and methylation character; Vanyushin BF et al.; The nucleotide composition, relative concentration of pyrimidine clusters, and the degree of methylation of the mitochondrial and nuclear DNA's of various vertebrates and the protozoan Crithidia oncopelti have been studied . With respect to the relative concentration of GC pairs, the mtDNA of animals (bull, rat) does not differ from the corresponding nDNA . The relative concentration of GC pairs in the mtDNA of certain fish and birds is 1.5-2.5 mole% higher than in the respective nDNA . The kinetoplast DNA of the protozoan C . oncopelti (where the relative concentration of the GC pairs is 42.9 mole %) differs very sharply in composition from the nDNA (where the relative concentration of GC pairs is 51.3 mole %) . The mtDNA's and kDNA's studied are distinguished from the respective nDNA'S by a lower degree of clustering of pyrimidine nucleotides . The proportion of mono- and dipyrimidine fragments in the mtDNA and kDNA is 30 mole %, while in the nDNA it does not exceed 23 mole % . The relative concentration of long pyrimidine clusters (hexapyrimidine clusters of larger) in the mtDNA is smaller than in the nDNA by a factor of 2-5 . The low degree of clustering of the pyrimidine nucleotides is apparently characteristic of all the known mtDNA's and may support the fact that they have a single type of organization and are of a single origin . All the vertebrate mtDNA's studied contain 5-methylcytosine as a minor base (1.5-3.15 mole %), and their level of methylation is 1.5-2 times greater than that in the respective nDNA's . It has been shown that animals display species specificity with respect to the 5-methylcytosine content in the mtDNA . Its distribution among the pyrimidine clusters in the bovine heart mtDNA differs substantially from that in the nDNA . This suggests that the methylation specificities of nuclear and mitochondrial DNA are different . A DNA methylase, which effects the in vitro methylation of cytosine residues both in the homologous mtDNA and in different heterologous DNA's, has been found in rat liver and bovine heart mitochondria . The specificity of the in vitro methylation of the cytosine residues in the same heterologous Escherichia coli B DNA by the nuclear and mitochondrial enzymes is different: The mitochondrial enzyme methylates predominantly in monopyrimidine fragments, and the nuclear enzyme methylates mostly in di- and tripyrimidine fragments . They, therefore, recognize different nucleotide sequences. Mol Biol (Mosk), 1976 Jul-Aug, 10(4), 682 - 5 Influence of antibodies against DNA on the renaturation of DNA; Ignashov VG; The treatment of denatured T4 phage DNA with antiserum for the DNA of this phage, containing antibodies against glucosylated 5-hydroxymethylcytosine, decreases the ability of DNA for renaturation . The greatest inhibiting activity is possessed by antiserum for T4 phage DNA irradiated with UV light, which contains antibodies not only against glucosylated 5-hydroxymethylcytosine, but also against the usual nitrogen bases . Antiserum against E . coli DNA, containing antibodies to the usual nitrogen bases, in equal dilutions with the antisera indicated above, shows less inhibitory activity on the renaturation of T4 phage DNA. Ateneo Parmense Acta Biomed, 1976 Jul-Aug, 47(4), 479 - 96 {Studies on the quality of water of the right side tributaries of the Po River; Trebbia and Nure, in 1971-1973}; Bellelli E et al.; In the period 1971-73 samplings were taken out monthly at the mouths of the two right side tributaries of the river Po, Trebbia and Nure . The aim of the research was to value the water quality, the polluted load conveyed to the river Po and to compare the real load with the one estimated on the basis of inhabitants and the basin area . Both the water courses are torrent-like rivers with highflow values in the winter and low flow values in the summer (July and August) . There are no important industries in the basins studied and the high polluted effluents, mainly domestic and agricultural discharges, are placed in the plain tracts of the rivers at few kilometers from the mouths . As far as the water quality of both rivers is concerned, most of the chemical parameters maintain suitable levels for fish life and drinking purposes; on the contrary the microniological indexes exceed the limits for bathing fixed by the Italian Ministry of Health by 100 E.coli 100/ml . and by 1000 total coliforms 100/ml . in about 90 and 70-80% of samplings . The comparison between real and calculated loads has shown a good agreement for BOD and chloride in Trebbia and for BOD and phosphates in Nure. Nucleic Acids Res, 1976 Jul, 3(7), 1671 - 87 Extensions of the known sequences at the 3' and 5' ends of 23S ribosomal RNA from Escherichia coli, possible base pairing between these 23S RNA regions and 16S ribosomal RNA; Branlant C et al.; Extensions of the known sequences at both 3' and 5' ends of 23S ribosomal RNA are presented: The 5' terminal is pG-G-U-U-A-A-G-Cp or pG-G-U.. . G-U-U-A-A-G-Cp, with a very short sequence between Up and Gp and the 3'terminal is G-A-A-C-C-G-A-(G)-G-C-U-U-A-A-C-C-U-UOH . These two terminal regions exhibit a high degree of complementarity . In addition, extensive complementarities are also found between the 5'terminal sequence of 23S RNA and a sequence contained in section A of the 16S ribosomal RNA, and between the 3'terminal sequence of 23S RNA and sequences in sections O and J in the 16S RNA . The degree of complementarity between the two extremities of 23S RNA, and between these extremities and regions of the 16S RNA, is far greater than would be expected on a random basis suggesting a possible involvement of this base-pairing in the functioning of ribosomes . This possibility is discussed. Nucleic Acids Res, 1976 Jul, 3(7), 1635 - 46 Location of protein S1 of Escherichia coli ribosomes at the 'A'-site of the codon binding site . Affinity labeling studies with a 3'-modified A-U-G analog; Pongs O et al.; An affinity analog with a 5-bromoacetamido uridine 5'-phosphate moiety bonded to the 3' end of A-U-G has been prepared with the aid of polynucleotide phosphorylase . This 3'-modified, chemically reactive A-U-G analog was used to probe the ribosomal codon binding site . The yield of the reaction depended strongly on the ribosomal source and was sensitive to salt-washing ribosomes . The major crosslinking product was identified to be protein S1 . Since the reaction of this 3'-modified A-U-G programmed ribosomes for Met-tRNA-Met-M binding, it is concluded that protein S1 is located at or near the 3'-side of the ribosomal codon binding site. Can J Biochem, 1976 Jul, 54(7), 650 - 6 Mechanisms of suppression in Drosophila . IV . Specificity and properties of tyrosyl-tRNA synthetase; Warner CK et al.; Tyrosyl-tRNA synthetase from wide type Drosophila can distinguish tRNATyr of wild type from that of the suppressor mutant, su(s)2 . The method of fractionation of the enzyme on DEAE-cellulose can produce three different forms of the enzyme . One of these forms is characterized in this study . We describe the apparent Km for K+, Mg2+, ATP, tyrosine and tRNA, as well as its heat stability and molecular weight . Spermine does not replace Mg2+ but is inhibitory, a Ki of 1 mM was obtained . Inhibition by p-chloromercuribenzoate results in a Ki of 1 muM . The properties of tyrosyl-tRNA synthetase of Drosophila are compared to the properties of this enzyme from E . coli, B . subtilis and yeast. Prikl Biokhim Mikrobiol, 1976 Jul-Aug, 12(4), 552 - 6 {Characteristics of asparaginase from Escherichia coli at different stages of purification}; Maksimov VI et al.; Asparaginase of Escherichia coli obtained according to the previously developed scheme (extraction, acidification, heating with ammonium sulphate, twice repeated precipitation with ethanol and DEAE-cellulose chromatography) was examined by the methods of polyacrylamide gel electrophoresis, isoelectric focusing and molecular weight assay . Asparaginase activity was measured by the Nessler method . With respect to the above characteristics, the asparaginase preparation was very similar to those described in the literature, e.g . Krasnitin (FRG) and Leunase (Japan). Mol Biol (Mosk), 1976 Jul-Aug, 10(4), 697 - 705 Use of the method of mixed substrates to study the specificity of tRNA methylases; Gambaryan AS et al.; The absence of summation of the rate of methylation of positionally analogous cytidine residues in tRNA1Val, tRNAPhe, and tRNAMet in the case of simultaneous presence of two substrates in the incubation mixture was demonstrated by the method of mixed substrates . The same result was also obtained in the methylation of A19 (counting from the 3' end of the molecule) in tRNA1Val, tRNAPhe, tRNAfMet, tRNASer, and tRNAGlu individually and in the case of their mixing in pairs . These data are evidence that positionally analogous nucleotides in different RNAs are attacked by the same enzyme . Yeast tRNASer, already possessing a methyl group at the cytidine residue studied, proved to be an effective inhibitor of methylase, forming m5C with valine and phenylalanine tRNAs . The results obtained are evidence that differences in the primary and secondary structures at the site of methylation are not the deciding factors in the interaction of tRNA with methylases. Mol Biol (Mosk), 1976 Jul-Aug, 10(4), 620 - 8 Isolation and study of some properties of the highly active 30S and 50S Escherichia coli ribosomal subunits; Semenkov YP et al.; A method for the isolation of highly active Escherichia coli ribosomal subunits has been described and used to obtain 30S subunits, which are fully active in the cistron-specific binding of tRNA, and reassociated 70S ribosomes, which are at least 35% active in the synthesis of polypeptides . The dissociation constants (Kd) of the 30S-poly(U)-tRNAPhe complex, which proved to be practically identical for tRNAPhe in the deacylated and aminoacylated forms, as well as for the chemically synthesized peptidyl-tRNA, have been measured . Changes in the binding conditions (temperatures from 0 to 30 degrees, Mg2+ concentrations from 20 to 5 mM, and NH4+ concentrations from 200 to 50mM) have a significant effect on the value of Kd without altering the number of active 30S subunits . It has been shown that the codon-specific binding of tRNA to the 30S subunits is completely reversible . The 30S subunits are not only not inactivated after a single act of binding of a tRNA molecule, but are capable of undergoing this process repeatedly without any appreciable loss in activity. Eur J Immunol, 1976 Jul, 6(7), 477 - 80 Reduction of trapping of aggregated human IgG in germinal centers by mitogens; Gutierrez C et al.; The effect of nonspecific mitogens on the trapping of 125I-labeled aggregated human IgG (125I-AHGG) in germinal centers (GC) of mouse spleens has been investigated by both radioactivity uptake and immunofluorescence . Phytohemagglutinin and concanavalin A (Con A) significantly decreased trapping . Lipopolysaccharide produced less inhibition, and pokeweed mitogen had no significant effect . The maximum inhibition occurred with 250--500 mug Con A . This had no effect on 125I-AHGG uptake in liver kidney and blood . No differences were found between i.p . and i.v . routes of Con A injection . The effect of mitogens on the 125I-AHGG trapping in GC is due more likely to modification of the migratory properties of lymphocytes brought about by surface binding, than to their mitogenic properties, since Con A decreased 125I-AHGG localization in thymectomized, x-irradiated and bone marrow-reconstituted animals. Nucleic Acids Res, 1976 Jul, 3(7), 1625 - 33 Comparison of the reactions of chemically reactive analogs of U-G-A and of A-U-G with ribosomes of Escherichia coli; Pongs O et al.; The chemically reactive analog of U-G-A, 5'-(4-(Bromo-{2-14C} acetamido) phenylphospho) - uridylyl-(3'-5') - guanylyl-(3'-5') adenosine has a 20 fold lower affinity to 70S ribosomes than the corresponding analog of A-U-G though the U-G-A analog also preferentially reacts with protein S18 of 70S ribosomes . This reaction programs ribosomes for EF-T dependent Trp-tRNATrp-suIII binding . Therefore, it is concluded that this protein is part of the A'-site of the ribosomal codon binding site . Reaction of the U-G-A analog with 30S subunits lead to a predominant crosslinking of U-G-A to proteins S4 and S18 . In contrast, a comparable reaction of the A-U-G analog with 30S subunits lead to a predominant crosslinking of A-U-G to proteins S4 and S12 (Pongs, O., Stoffler, G.A., Lanka, E., (1975) J . Mol . Biol . 99, 301) . Since protein S12 is located at the 'P' site of the ribosomal codon binding site, it is proposed that the U-G-A analog does not bind at this site. Pahlavi Med J, 1976 Jul, 7(3), 371 - 86 Colonic malakoplakia in a child: report of a case and review of literature; Taghinia MA et al.; The fifth case of colonic malakoplakia involving children under the age of 13 years is reported . Rectoscopic findings were strongly suggestive of a colonic carcinoma . A detailed review of literature on malakoplakia is also made. Can J Microbiol, 1976 Jul, 22(7), 922 - 8 The conversion of leucine to alpha-ketoisocaproic acid and its metabolic consequences for Escherichia coli K12; Newman EB et al.; The amino acid L-leucine serves as a good auxiliary nitrogen source for Escherichia coli K12, and in so doing is converted to alpha-ketoisocaproic acid which is excreted into the medium . L-Leucine does not serve as sole nitrogen source . Cells incubated with L-leucine as sole nitrogen source do not grow, although they do metabolize leucine, and accumulate ketoisocaproic acid in the medium . Where glycine is the only other nitrogen source, the presence of L-leucine greatly increases the growth rate even at concentrations so low that its contribution as nitrogen donor is unlikely to be important. Biophys Chem, 1976 Jul, 5(1-2), 271 - 83 The so20,w of unsheared DNA from whole cell lysates of Escherichia coli; Appleby DW et al.; We present measurements of the sedimentation coefficients of DNA present in whole cell lysates of E . coli . The method used is a preparative version of the band sedimentation experiment of Bruner and Vinograd . We show that in order to obtain reliable data on the time dependence of sedimentation, it is necessary to accelerate and decelerate the rotor over much longer times than the standard centrifuge allows . We describe the necessary modifications to the preparative centrifuge and use them to determine the So20,W of unsheared E . coli DNA . The value for the fastest moving components in the lysate is 220 S . The molecular weight of the DNA corresponding to this sedimentation coefficient is probably 1.7 X 10(9) g/mole . However, alternative values cannot be ruled out. Biochem J, 1976 Jul 1, 157(1), 221 - 7 Replication of the deoxyribonucleic acid of multiple-drug-resistance factor in Escherichia coli; Barker GR et al.; 1 . It was shown that a system previously described for labelling R-factor DNA during transfer to an irradiated recipient strain of Escherichia coli did not allow high selectivity in the incorporation of thymine into R-factor DNA . 2 . Lack of selectivity was shown to be due to cross-feeding from recipient to donor strain . 3 . An improved system using a nalidixic acid-resistant recipient strain is described in which incorporation of thymine into the DNA of donor cells is minimized by addition of nalidixic acid after completion of transfer of the plasmid during conjugation. Biochem J, 1976 Jul 1, 157(1), 207 - 10 The specificity of a 7 alpha-hydroxy steroid dehydrogenase from Escherichia coli; Haslewood ES et al.; 1 . Thirty-eight steroids were tested as substrates for a 7 alpha-hydroxy steroid dehydrogenase preparation from a strain of Escherichia coli; an improved method of making the crude enzyme is described . 2 . Steroids having a 7 alpha-hydroxyl group in the molecule were substrates except (a) when the 5 beta-cholan-24-oic acid side chain was shortened to less than four carbon atoms and (b) in certain cases when sulphate ester groups were present in the molecule . 3 . For testing with the enzyme, a new specimen of 7 alpha-hydroxy-3,12-dioxo-5 beta-cholan-24-oic acid was made, which had properties different from those previously described. Am Ind Hyg Assoc J, 1976 Jul, 37(7), 427 - 31 A method for biological testing of containment systems for viral agents; Bolton NE et al.; A technique utilizing coliphage as the test material has been developed and employed to evaluate the effectiveness of a containment system for zonal centrifugation of hepatits viruses . An Andersen Viable Particle Sampler which had been loaded with plates containing a base layer of agar nutrient with an overlay of E . coli- agar suspension was used to sample the test air . The containment system, which includes a HEPA filter, was challenged with an aerosolized suspension of coliphage. Mutat Res, 1976 Jul, 40(3), 229 - 35 Mutagenic actions of chlorocholine chloride; Sussmuth R et al.; Chlorocholine chloride, at concentrations of 0.2 to 0.5 M (3.2 to 7.9%) between pH 5 and 8, showed no significant mutagenic effect whereas at the same concentrations at pH 9 it caused mutations, in a valine-sensitive strain of E . coli K12 . Treatment of E . coli B with chlorocholine chloride at 0.5 M and pH 9 resulted in auxotrophic and regulatory deficient (valine-sensitive) mutants . The mutagenic effects of chlorocholine chloride were compared with the effects caused by the food additive NaHSO3. J Clin Microbiol, 1976 Jul, 4(1), 82 - 6 Antigen distribution in a latex suspension and its relationship to test sensitivity; Hechemy K et al.; Specific aggregation of latex particles due to antigen-antibody reaction occurred only when antigen was latex bound, as found in a study using Escherichia coli 0111:B4 extract . Maximum serological sensitivity was achieved when free antigen was present in a narrow range of ratios with the bound antigen. J Chromatogr Sci, 1976 Jul, 14(7), 350 - 3 A method for the rapid separation and characterization of biological pyridines; Averett DR et al.; Pyridine compounds of biological origin are separable by 2-dimensional descending paper chromatography . When assayed in situ for a tertiary or quaternary ring nitrogen, each compound gives a charactertistic reaction which is not predictable from the structural configuration . A cell-free extract of Escherichia coli, incubated in the presence of isoniazid, yields an ultraviolet light quenching spot with Rf values different from any tested standard . The compound is hypothesized to be a metabolite of either nicotinic acid or isonazid. J Antibiot (Tokyo), 1976 Jul, 29(7), 754 - 8 Inhibition of coliphage multiplication and R plasmid transfer by desdanine; Tanida S et al.; Desdanine inhibited the plaque formation of male-specific coliphages but not that of other coliphages tested . Desdanine also suppressed the multiplication of both RNA phage Q beta and filamentous DNA phage f1 at the concentration of 3.13 approximately 6.25 mug/ml which had no influence on the growth of their host cells . However, the inhibitory effect on the phage multiplication was not due to the inactivation of phage particles nor the prevention of phage adsorption and penetration into the host cells . Desdanine also inhibited the transfer of R plasmid, R 100-1, in E . coli at 6.25 approximately 12.5 mug/ml without affecting the viability of donor and recipient cells. Am J Surg, 1976 Jul, 132(1), 59 - 63 Prevention of wound infections . A case for closed suction drainage to remove wound fluids deficient in opsonic proteins; Alexander JW et al.; Fluids collecting in surgical wounds in both dogs and man have been shown to lose progressively the ability to opsonize bacteria for phagocytosis and killing of bacteria by normal neutrophils . Since the collection of fluids in potentially contaminated wounds also interferes with access of phagocytic cells to contaminating bacteria and provides a pablum for growth, their removal seems to be indicated to minimize the risk of infection . This can be accomplished easily and safely with the use of closed suction drainage as demonstrated in 100 patients undergoing bilateral nephrectomy, splenectomy, and renal transplantation. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2379 - 83 New low resolution model for 50S subunit of Escherichia coli ribosomes; Stuhrmann HB et al.; Neutron low angle scattering studies of the 50S subunit of E . coli ribosomes with the contrast variation method reveals large fluctuations in the scattering density . A region of relatively low scattering density, rich in proteins, surrounds an RNA-rich core of higher scattering density . The centers of mass of the RNA and protein parts of the 50S subunit are separated by a distance (20 A) that is considerably smaller than that reported in previous studies. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2316 - 20 Isolation of inverted repeat sequences, including IS1, IS2, and IS3, in Escherichia coli plasmids; Ohtsubo H et al.; A method is described for isolation of inverted repeat DNA sequences that occur in E . coli plasmids . The procedures of the isolation involved: (a) denaturation of intact plasmid DNA, (b) a rapid, 30 sec, renaturation of inverted-repeat sequences in the genome, (c) digestion of the single-stranded portion by S1 nuclease to recover duplex DNA, and (d) detection and purification of the duplexes using 1.4% agarose gel electrophoresis . If a plasmid DNA carried inverted repeats of either one type or two different types of special DNA sequences, these procedures enabled us to observe either one or two characteristic DNA bands, respectively, in the agarose gels . If a plasmid DNA did not carry any inverted repeats, or if the plasmid DNA only carried direct repeat sequences, no characteristic DNA bands were recovered . Cleavage of the spacer DNA between inverted repeat sequences generated no gel bands . This indicated that the inverted repeat sequences must be in the same strand . Using this method, we isolated and purified several repeated sequences, including IS1, IS2, and IS3, from derivatives of F and R plasmids. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2285 - 8 Regulation of transcription factor rho and the alpha subunit of RNA polymerase in Escherichia coli B/r; Blumenthal RM et al.; Transcriptional termination factor rho, the alpha subunit of RNA polymerase (RNA nucleotidyltransferase nucleosidetriphosphate: RNA nucleotidyltransferase, EC 2.7.7.6), and ribosomal protein S6 were resolved from whole-cell extracts of E . coli B/r by a high-resolution, two-dimensional polyacrylamide gel electrophoretic technique, and were identified through coelectrophoresis with the purified proteins . The regulation of rho, alpha, and S6 was studied, in steady-state cultures of E . coli B/r growing at rates ranging from 0.6 to 2.1 generations per hr, through the use of this gel technique and a double radioisotope labeling procedure . The regulatory patterns of rho and alpha are distinct from, but similar to, one another . Neither rho nor alpha shows the sharply increasing levels with increasing growth rate shown by the ribosomal proteins was exemplified by S6 . The difference between the levels of rho and alpha, on the one hand, and S6, on the other, is most pronounced during rapid growth . The regulatory pattern of alpha is interesting, given the recent suggestion that the gene coding for alpha is contranscribed with genes coding for ribosomal proteins. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2221 - 5 Role of 5S RNA in assembly and function of the 50S subunit from Escherichia coli; Dohme F et al.; Total reconstitution experiments performed under various conditions revealed that 5S RNA plays an important role during the last assembly step in vitro leading to an active 50S particle . For the preceding steps this RNA species is dispensable . However, 50S RNA can be integrated efficiently during any of the assembly steps in vitro . The 47S particle, reconstituted in two steps and lacking 5S RNA, shows low but significant activity in many functional tests . High activity could be obtained by incubating this particle with 5S RNA alone, demonstrating the importance of the 5S RNA in generating an active ribosomal conformation . In particular, the activity of the peptidyltransferase (peptidyl-tRNA:aminoacyl-tRNA N-peptidyltransferase; EC 2.3.2.12) center is drastically influenced by 5S RNA . No significant factor-dependent tRNA binding to the A-site was observed with the 47S particle, in contrast to the corresponding P-site binding . The elongation factor G dependent GTPase activity was not affected by the lack of 5S RNA. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2216 - 20 Physical studies of denatured tRNA2Glu from Escherichia coli; Bina-Stein M et al.; We have examined the 270 MHz nuclear magnetic resonance spectrum and relaxation kinetic behavior of tRNA2Glu (E . coli) in the absence of Mg++, a condition which produces an inactive form of this tRNA . The results show that the denatured form has about five fewer proton resonances in the region from -12 to -15 ppm . Relaxation kinetic measurements reveal that the denatured conformer contains three separately melting helices . The results support a model in which the tertiary structure and dihydrouridine helix characteristic of the native form are unfolded in the denatured state, and are replaced by an altered tertiary structure . The acceptor stem, anticodon, and TpsiC helices are intact in this model for the denatured conformation . The optical changes that accompany melting of the denatured tertiary structure are faster than 10 musec. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2186 - 90 Structure of L-arabinose-binding protein from Escherichia coli at 5 A resolution and preliminary results at 3.5 A; Phillips GN Jr et al.; The three-dimensional crystal structure of the L-arabinose-binding protein from E . coli, an essential component in the active transport of L-arabinose, has been solved at 5 A resolution using the method of multiple isomorphous replacement . Five heavy atom derivatives were used . A preliminary 3.5 A electron density map has also been calculated . The results indicate that the molecule is ellipsoidal with approximate dimensions 68 A X 38 A X 30 A . Two similar domains within the molecule (which is a single polypeptide chain) are related by an approximate noncrystallographic rotation-translation axis . This relationship involves approximately 20% of the structure. Mutat Res, 1976 Jul, 36(1), 17 - 28 Partial suppression of the LexA phenotype by mutations (rnm) which restore ultraviolet resistance but not ultraviolet mutability to Escherichia coli B/r uvr A lexA; Volkert MR et al.; In Escherichia coli, lexA mutations eliminate expression of UV-inducible functions, causing pleiotropic effects which include sensitivity to ultraviolet (UV) light and loss of UV mutability . Selection for UV resistance, after 5-bromouracil (BU) treatment of E . coli B/r uvrA lexA-102, has yielded derivatives more resistant than lexA but still refractory to UV mutagenesis . The mutation responsible for the UV-resistant UV-nonmutable phenotype (rnm) is cotransducible with malB to about the same extent as is lexA-102 and is tightly linked to lexA-102 in at least one strain . The rnm mutation may therefore be an intragenic partial suppressor of the LexA phenotype . In addition to increased UV resistance and lack of UV mutability, rnm strains show improved ability to perform postreplication repair and to control postirradiation DNA degration compared to the lexA parent . We ascribe the properties of rnm mutants to their having reacquired control of Exonuclease V activity without having reacquired UV-inducible error-prone postreplication repair . We relate our results to current interpretations of UV mutagenesis and to models of coordinate regulation of UV-inducible functions. Mutat Res, 1976 Jul, 36(1), 11 - 6 Formaldehyde induced DNA-protein crosslinks in Escherichia Coli; Wilkins RJ et al.; Exposure of Escherichia coli to low doses of formaldehyde induces interstrand cross-links in the cellular DNA, at least 50% of which involve protein "bridges" between the DNA strands . The biological importance of these cross-links is suggested by both the high yield of formation and by the inability of some sensitive repair deficient mutants to completely remove cross-links and bound protein from the DNA during post treatment incubation. J Virol, 1976 Jul, 19(1), 271 - 4 Genetic studies of coliphage P1 . II . Relatedness to P7; Walker DH Jr et al.; Using semiquantitative spot tests, 107 independently isolated amber mutants of P1 were shown to be rescued by a nonpermissive strain of Escherichia coli lysogenic for P7 (previously called phiamp), indicating extensive genetic relatedness between P1 and P7 . The amount of rescue observed varied with mutants from different genetic linkage clusters of P1 . Although these rescue tests cannot distinguish between recombination, complementation, transactivation, or combinations thereof, a major role is indicated for recombination. J Med Chem, 1976 Jul, 19(7), 903 - 8 Synthetic inhibitors of Escherichia coli, calf thymus, and Ehrlich ascites tumor thymidylate synthetase; Kampf A et al.; In a study of active site binding the inhibition of thymidylate synthetase derived from Escherichia coli, calf thymus, and Ehrlich ascites tumor was examined using eight inhibitors . 5-Substituted 2'-deoxyuridine 5'-phosphate analogues used in this study are the hydroxymethyl, methoxymethyl, benzyloxymethyl, formyl, acetyl, allyl, and two potential active site alkylating substituents: 2,3-oxypropyl and the azidomethyl analogues . All compounds were competitive with the substrate, 2'-deoxyuridine 5'-phosphate; the most potent inhibitor was 5-formyl-dUMP (Ki = 0.1, 0.09, and 0.08 muM for the respective enzyme) . The 5-hydroxymethyl, 5-benzyloxymethyl, and 5-azidomethyl derivatives of dUMP showed some differential inhibition; these compounds were two to three times more active against the ascites tumor enzyme than against the thymus enzyme. Infect Immun, 1976 Jul, 14(1), 95 - 9 Suckling mouse model for detection of heat-stable Escherichia coli enterotoxin: characteristics of the model; Giannella RA; Although the suckling mouse assay is widely used for the detection of heat-stable Escherichia coli enterotoxin (ST), few data have been published concerning the reproducibility, optimal growth, and test conditions of this assay . Four strains of toxigenic E . coli known to elaborate both heat-labile enterotoxin and ST or ST alone were used to study these parameters . ST activity after heat treatment and the effect of purified choleragen were also examined . ST production was optimal in Casamino Acids-yeast extract media, but both Trypticase soy and brain heart infusion broths resulted in several false negative reactions . Growing cultures in roller tubes was the most reliable method of ST production . Shaking-flask cultures and stationary-grown cultures resulted in suboptimal ST production in several strains . Optimal mouse incubation time was 3 h, and fluid secretion did not rise thereafter . Adequate toxin production occurred after 16 to 24 h of incubation . The coefficient of variation of various toxins tested on many occasions varied between 10.5 and 15.7% . Toxin activity was stable for 6 months when frozen at - 20 C . There was no decrease in ST activity when heated at 65 C for 15 min, but a small decrease was observed in two of four strains after heating at 100 C for 30 min . Choleragen, tested at various doses and at multiple times, gave uniformly negative results . These studies indicate that when done under the proper conditions, the suckling mouse assay is a simple, rapid, and reproducible assay for E . coli ST. Eur J Biochem, 1976 Jul 1, 66(2), 369 - 77 Cell-wall lipopolysaccharides of ampicillin-resistant mutants of Escherichia coli K-12; Prehm P et al.; The lipopolysaccharides of ampicillin-resistant cell-wall-defective mutants of Escherichia coli K-12 were analyzed . From their lipopolysaccharides the respective core oligosaccharides were obtained . Following dephosphorylation,the core oligosaccharides were methylated and analyzed by gas chromatography/mass spectrometry . From core-defective mutants substructures of the K-12 core were obtained . Analysis of the lipopolysaccharide preparations from wild-type K-12 indicated the presence of several core structures with different degrees of completion . The lipopolysaccharide preparation was degraded and the oligosaccharide mixture was partially resolved by gel filtration chromatography . Methylation, gas chromatography and mass spectrometry of the oligosaccharides permitted the tentative formulation of the K-12 core structure . Alternative interpretations for this heterogeneity are discussed. Eur J Biochem, 1976 Jul 1, 66(2), 357 - 68 Immunochemical studies on lipopolysaccharides from wild-type and mutants of Escherichia coli K-12; Mayer H et al.; Lipopolysaccharides from a number of mutants of Escherichia coli K-12 were investigated by means of chemical and serological methods . Inhibition of passive hemagglutination and inhibition of precipitation show that L-rhamnose is the immunodominant sugar in the lipopolysaccharide from wild-type E . coli K-12 . The disaccharide rhamnosyl-KDO (where KDO is 3-deoxy-D-manno-octulosonic acid) was isolated and characterized after mild acid hydrolysis of the lipopolysaccharide . It is concluded that rhamnose is present in the innermost part of the core as a side-chain substituent on KDO . From crosses between an E . coli K-12 donor and E . coli O8, hybrids were obtained which contained either one or both of the donor rfa and rfb clusters . Serum absorption studies with lipopolysaccharides from these hybrids indicated that the histidine-linked rfb cluster is responsible for the presence of rhamnose in the K-12 core oligosaccharide . Using paper chromatography of 32P-labelled lipopolysaccharides we have found heterogeneous lipopolysaccharide in two strains as well as some differences between two wild-type strains . The latter difference is believed to be due to varying contents of KDO-linked ethanolamine phosphate . The overall results presented together with those described in the companion paper clearly show that the core oligosaccharide in E . coli K-12 has a structure different from the types previously described for other strains of E . coli (designed coli R1 to coli R4). Arch Surg, 1976 Jul, 111(7), 783 - 6 Total body washout for the treatment of endotoxin shock . An experimental study; Nakamura Y et al.; In order to study the therapeutic effects of total body washout (TBW) in experimental endotoxin shock, we used the following procedure . Seventeen rabbits (controls) received Escherichia coli endotoxin (5 mg/kg) intravenously and were observed for 12 hours . Shock developed in 14 rabbits; they died in 5.2 +/- 1.0 (mean +/- SD) hours, with a survival rate of 18% . Seventeen rabbits were subjected to TBW only . Muscle temperature was lowered to 25 C with a pump oxygenator circuit and the animals were exsanguinated . After residual blood was flushed out with cold, lactated Ringer solution, the animals were rewarmed with another circuit that was primed with homologous blood . Fourteen animals survived (82%) . Two hours after E . coli endotoxin was injected intravenously 17 animals were treated with TBW . The survival rate (53%) of this group was significantly higher than in the control group (18%) (P less than .005) . Eight nonsurvivors showed hypotension and acidosis even after TBW treatment, thus indicating the irreversibility of their endotoxin shock . This study indicates that endotoxin shock may be reversed by TBW if it is instituted before irreversible cellular damage. Am J Vet Res, 1976 Jul, 37(7), 831 - 4 Vaccination of cows with an Escherichia coli bacterin for the prevention of naturally occurring diarrheal disease in their calves; Myers LL; A formalin-killed Escherichia coli bacterin composed of 6 enterotoxigenic strains of the organism prepared from calves with diarrheal disease with field tested for efficacy against naturally occurring diarrheal disease in young calves . The bacterin was tested in 23 privately owned beef herds in Montana involving 3,508 cows and their calves . About half the cows in each herd were given 2 subcutaneous vaccinations before calving and the other half (controls) were injected twice with a placebo . Almost all of the participating herds had a record of acute diarrheal disease . The number of calves from vaccinated dams that died of diarrheal disease was significantly less (P = 0.004) than the number of calves from control dams that died of diarrheal disease (60 vs 99 calves) . There was no significant difference in the number of calves in the groups that developed mild diarrhea . There was marked interherd variation in vaccinal efficacy, possibly due in part to differences in the cause of diarrheal disease between herds . Colostral agglutinating antibody titers against formolized cells of each of the 6 vaccinal strains of E coli were markedly higher in vaccinated dams than in control dams . The agglutinating antibody titers were highest against enterotoxigenic E . coli strains 4 and 5 and lowest against strains 2 and 6 . Presumably, colostral antibody passively transferred to the calf at the time of nursing was responsible for the protection observed. Acta Paediatr Scand, 1976 Jul, 65(4), 417 - 23 Escherichia coli O antibody content in milk from healthy Swedish mothers and mothers from a very low socio-economic group of a developing country; Carlsson B et al.; The antibody content of milk from healthy Swedish mothers was compared with that of milk from mothers of a very low socio-economic group in a developing country . Antibodies of various immunoglobulin classes against E . coli O antigens were determined with the enzyme-linked immunosorbent assay (ELISA) . The milk antibodies which mainly belonged to the secretory IgA class appeared in similar concentrations in milk from the two groups using E . coli antigens of Swedish as well as Pakistani origin . The secretory IgA antibodies could be demonstrated in the stool of the breast-fed infants of the undernourished mothers . Also the concentration of serum IgG and IgA antibodies to E . coli O antigens were similar in the Pakistani and Swedish mothers . The serum IgM antibody levels of the Pakistani mothers were higher, however, presumably due to a higher frequency of infections . It was noted that the milk production decreased considerably upon the hospitalization of the healthy and well-to-do Swedish mothers . The small milk volumes of the undernourished Pakistani mothers suggest that the lactation failure observed was mainly due to inadequate milk flow and not to decreased milk quality . The results indicate the necessity of studying the nutritional, psychological and social factors responsible for low milk yield and add yet another reason to stimulate prolonged breastfeeding. Obstet Gynecol, 1976 Jul, 48(1), 31 - 4 The predictability of intrauterine infection by analysis of amniotic fluid; Listwa HM et al.; Amniotic fluid samples from 95 internally monitored patients were examined to determine whether the appearance of polymorphonuclear leukocytes or bacteria could predict intrauterine infection . All patients delivered vaginally . More than one polymorphonuclear leukocyte per oil field were seen in specimens of 32% of patients; bacteria were seen in specimens of 52% of patients, and organisms were grown in 93% of specimens, yet, the overall maternal infection rate was only 6.3% . Moreover, infection developed in only 10% of patients with polymorphonuclear leukocytes in the fluid and 6% of patients with positive gram stain or cultures . For patients who deliver vaginally, the appearance of polymorphonuclear leukocytes or bacteria does not predict infection. J Thorac Cardiovasc Surg, 1976 Jul, 72(1), 150 - 6 Long-term follow-up of aortic valve replacement with the fresh aortic homograft; Anderson ET et al.; The long-term results of aortic valve replacement with the fresh aortic homograft, performed in 114 patients at Stanford University Medical Center from 1967 to 1971, were evaluated . There were 10 operative deaths (8.8 per cent), only 3 (5 per cent) in the period from 1968 to 1971 . There were 6 late deaths in the first year (5.8 per cent) and 8 in later years (1.5 per cent per year); 12 late deaths were due to cardiac causes, 6 of them to valve dysfunction . The homograft was replaced later with a prosthetic valve or heterograft in 22 patients (3.2 per cent per year): for regurgitation in 20 and for calcific stenosis in only one . Infective endocarditis occurred in 5 cases, accounting for one operative death, 2 late deaths, and 2 reoperations with survival . Systemic thromboembolism occurred in 6 patients, 3 with mitral valve disease, one with atrial fibrillation, and one with infective endocarditis; none was a proved instance of embolism from bland thrombus on the aortic homograft valve . Of 53 patients followed for 5 years or more with the homograft intact, 47 have minimal or no disability, despite aortic diastolic murmurs in many . We conclude that long-term results are good in the majority of patients, with aortic regurgitation requiring reoperation being the leading complication . These results may serve as a basis for comparison of more recently introduced methods of aortic valve replacement. J Pediatr, 1976 Jul, 89(1), 8 - 10 Enterotoxigenicity and invasive capacity of "enteropathogenic" serotypes of Escherichia coli; Echeverria PD et al.; Forty-two strains of Escherichia coli that agglutinated in pools of antisera used to identify "enteropathogenic" serotypes were tested for heat-labile and heat-stable toxin production and for their ability to invade intestinal mucosa . None of the strains tested were enterotoxigenic or enteroinvasive as determined by the adrenal cell (heat-labile toxin), the suckling mouse (heat-stable toxin), or guinea pig eye (invasive capacity) assays . Our observations suggest that serotyping of E . coli is an unreliable method to identify isolates that are capable of causing gastroenteritis, at least as determined by available in vitro techniques. J Bacteriol, 1976 Jul, 127(1), 76 - 83 Selection for citrate synthase deficiency in icd mutants of Escherichia coli; Lakshmi TM et al.; Cultures of isocitrate dehydrogenase-deficient (icd) mutants were overgrown by double mutants (icd glt) lacking citrate synthase activity also . The icd mutants grew more slowly than wild-type cells or the double mutants because they accumulated an inhibitory metabolite (possibly citrate) . Intracellular citrate levels were several hundred-fold higher in icd cells than in wild-type or icd glt cells . Final growth yields of the wild type and the icd mutant on limiting glucose were equivalent and greater than the growth yield of icd glt double mutants . The icd gene mapped between 60 and 74 min . icd mutants were resistant to nalidixic acid, but glt and icd glt mutants and wild-type cells were sensitive, indicating that resistance results from accumulation of isocitrate, citrate, or a derivative of these compounds. J Bacteriol, 1976 Jul, 127(1), 656 - 9 Co-regulation of the phosphate-binding protein and alkaline phosphatase synthesis in Escherichia coli; Yagil E et al.; In phosphate-starved cells of Escherichia coli, the synthesis of alkaline phosphatase and some additional periplasmic proteins is derepressed . One of these proteins, which does not appear in a phoS- constitutive strain, has been identified as well the periplasmic phosphate-binding protein. J Bacteriol, 1976 Jul, 127(1), 564 - 71 Ultrastructure of paracrystals of a lipoprotein from the outer membrane of Escherichia coli; DeMartini M et al.; The highly purified lipoprotein of the outer membrane of Escherichia coli forms paracrystals . The ultrastructures of these paracrystals were examined by electron microscopy . The needle-shaped paracrystals show several different band patterns, depending on conditions of paracrystallization . Models are presented to explain possible arrangements of the lipoprotein molecules within the paracrystals. J Bacteriol, 1976 Jul, 127(1), 46 - 50 R factor-mediated polarized chromosomal transfer in Escherichia coli C; Heden LO et al.; Five transferable drug resistance factors (R factors) with the ability to bring about chromosomal transfer in Escherichia coli C were investigated with respect to the direction and origin of chromosome transfer . They were found to constitute two groups, both with a clockwise direction of transfer . One group has its origin of transfer between arg and pro, and the other group has its origin of transfer between try and man on the E . coli C chromosome . With one R factor in particular, the rapid increase in the production of recombinants was followed by a decrease in numbers as mating was prolonged. J Bacteriol, 1976 Jul, 127(1), 418 - 32 Regulation of beta-glucuronidase synthesis in Escherichia coli K-12: pleiotropic constitutive mutations affecting uxu and uidA expression; Novel M et al.; Among the beta-glucuronidase (UID)-constitutive mutants obtained by growth on methyl-beta-D-galacturonide, some strains are also derepressed for the two enzymes of the uxu operon: mannonate oxidoreductase (MOR) and mannonate hydrolyase (HLM) . By conjugation and transduction experiments, two distinct constitutive mutations were separated in each pleiotropic mutant strain . One of them was specific for uidA gene expression and was characterized as affecting either uidO or uidR sites . The second type of mutation was mapped close to the uxu operon and was found to be responsible for the pleiotropic effect revealed in the primary mutants: after separation such a mutation still fully derepresses MOR and HLM synthesis but weakly derepresses UID synthesis . The pleiotropic effect of this mutation was maintained even though the activity of the structural genes was altered . This rules out the occurrence of an internal derepressing interaction between these enzymes . In merodiploid strains, uxu-linked constitutive mutations were recessive to the wild-type allele, suggesting that these mutations could affect a regulatory gene . The uxuR gene is probably a specific regulatory gene for a very close operon, uxu . Moreover, it has a weak effect on uidA expression . Thus, UID synthesis would be negatively controlled through the activity of two repressor molecules that are synthesized by two distinct regulatory genes, uidR and uxuR . These two repressing factors are antagonized, respectively, by phenyl-thio-beta-D-glucuronide and mannonic amide and could cooperate in a unique repression/induction control over uidA expression . Constitutive mutations affecting the control sites of uidA gene probably characterize two distinct attachment sites in the operator locus for each of the repressor molecules. J Bacteriol, 1976 Jul, 127(1), 406 - 17 Regulation of beta-glucuronidase synthesis in Escherichia coli K-12: constitutive mutants specifically derepressed for uidA expression; Novel M et al.; All methyl-beta-D-galacturonide-positive mutants isolated from Escherichia coli K-12 carry constitutive mutations for beta-glucuronidase (UID) synthesis . Most of these mutants are specific for UID synthesis and are distributed in three classes according to the derepression level of UID . Each specific mutant carries a mutation(s) near uidA, the structural gene for UID, at min 30.5 of the E . coli K-12 linkage map . The expression of UID synthesis in F-merodiploid strains carrying these mutations permits discrimination between dominant and recessive constitutivity over the wild-type allele . The first kind of mutation (dominant) should affect the operator site uidO of the structural gene uidA; the second type of mutation (recessive) should affect a regulatory gene, uidR, operating through a negative control . The isolation of mutants bearing at this locus superrepressed mutations, which can revert to produce a constitutive phenotype, confirms the occurrence of such a regulatory gene . The partially derepressed uidR mutants of the first class are normally inducible and remain constitutive at low temperature; their UID has the same thermal sensitivity as in the wild-type strains . The occurrence of similar regulatory gene mutants has been recently described in the lactose system (Shineberg, 1974). J Bacteriol, 1976 Jul, 127(1), 348 - 53 Expression of a mutation affecting F incompatibility in the integrated but not the autonomous state of F; Pfister A et al.; Previously we have described a mutant Hfr strain in which incompatibility between the integrated F factor and an autonomous F-prime (F') factor was abolished . The mutation (inc) was located in the integrated F factor . F-prime factors isolated from the mutant Hfr strain have the same incompatibility behavior as those isolated from normal Hfr strains . Reintegration of these F' factors into the chromosome restores the Inc- phenotype characteristic of the mutant Hfr . The inc mutation thus affects incompatibility between integrated F and autonomous F(Fi-Fa incompatibility) but not incompatibility between two autonomous F factors (Fa-Fa incompatibility) . The implications of this finding for the mechanism of plasmid incompatibility are discussed. J Bacteriol, 1976 Jul, 127(1), 32 - 9 Wild-type and mutant in vitro products of an operon for ribonucleic acid polymerase subunits; Austin S; An in vitro protein-synthesizing system can synthesize two ribonucleic acid (RNA) polymerase subunits of Escherichia coli, beta and beta', when a transducing phage deoxyribonucleic acid (DNA) template containing the rpoB region of the bacterial chromosome is added . Recombinant rpoB transducing phages were isolated that carry "nonsense" mutations of the class rpo-rifampin zero amber (formally referred to as rifoam) . DNA was extracted from two of these phages . These DNAs are unable to direct the synthesis of beta subunits, whereas beta' synthesis is unaffected . Both mutations can be efficiently suppressed in vitro by the addition of suppressor transfer RNA . One of the mutations (rpoB115) produces a detectable nonsense (or restart) fragment of the beta protein in the absence of suppression . It is concluded that rpoB115 is an amber mutation within the structural gene for the beta subunit of RNA polymerase. Fed Proc, 1976 Jul, 35(9), 2026 - 30 Plasmid DNA replication; Helinski DR; Recent studies have provided some insight to the overall characteristics of plasmid replication in bacteria . The ColE 1 and R6K plasmids replicate via a covalently-closed circular intermediate . Replication is initiated at a fixed origin and is unidirectional in the case of ColE 1 and bidirectional for R6K . In the case of the plasmid R6K . the bidirectional replication is asymmetric and sequential, proceeding from a fixed origin to a fixed terminus located approximately 20% of the R6K genome from the origin . RNA serves as a primer for plasmid DNA replication . The base composition and sequence of the 5'-terminus of the RNA segments in ColE 1 DNA have been determined . Finally, the properties of plasmid relaxation complexes and the possible role of these complexes in plasmid DNA replication are discussed. J Biochem (Tokyo), 1976 Jul, 80(1), 111 - 20 Ca2+-induced conformational changes of spin-labeled g2 chain bound to myosin and the effect of phosphorylation; Okamoto Y et al.; One of the low molecular weight components of myosin, g2, was isolated by alkali treatment of myosin and was chemically modified with a spin label reagent, 4-maleimido-2,2,6,6-tetramethylpiperidinooxyl . The label on g2 showed a rather weakly immobilized ESR spectrum and it was clearly affected by Ca2+; the half-maximal change was at around pCa 4 . The spin-labeled g2 was incorporated into myosin by exchange with the intrinsic g2 of myosin in 0.6 M KSCN or 4 M LiC1 . The label on g2 became strongly immobilized on association with myosin . Under the conditions used, ESR spectral change due to Ca2+ occurred at two different concentration ranges, which were as low as pCa 8 and at around pCa 4 . Phosphorylated g2 was isolated from myosin after the protein kinase {EC 2.1.1.37}-catalyzed phosphorylation of myosin and it was also modified with the maleimide label . Dephosphorylation of the phosphorylated g2 was performed using E . coli alkaline phosphatase {EC 3.1.3.1} . The effects of Ca2+ on the ESR spectra of phosphorylated and dephosphorylated g2 were investigated on the state associated with myosin . A change in the ESR spectrum from strongly immobilized to weakly immobilized states was observed with both g2 chains on the addition of Ca2+ . However, the effective concentration ranges of Ca2+ were quite different; around pCa 4 for the phosphorylated g2 and around pCa 8 for the dephosphorylated g2 . The results indicate that g2 undergoes a conformational change at physiological levels of Ca2+ sufficient to saturate troponin, but it does not do so after phosphorylation. Nord Vet Med, 1976 Jul-Aug, 28(7-8), 368 - 76 {The influence of infectious bronchitis virus on egg production, fertility, hatchability and mortality rate in chickens (author's transl)}; Bisgaard M; In 35 flocks comprising almost 100,000 White Plymouth Rock birds producing hatching eggs, infectious bronchitis (IB) caused a decline in production which appeared to be connected with the age of the birds, when the outbreak started (table I) . In outbreaks of IB in pullets 3-4 weeks before they are ready to lay there is a considerable risk of delay of production and a generally too low yield (fig 1) . Egg laying birds, 25-34 weeks of age, suffered a drop in production from 20-80% (on an average 40%) and normal yield was never regained . After 6-8 weeks the production rate usually stabilizes around 6-12% below normal production . IB outbreaks in the middle of the production period (35-45 weeks) resulted in a decline of egg production of 40-90% (on an average 55%) . Recovery of production usually occurred after 6-8 weeks (fig . 2) . Observations of clinically typical outbreaks without any laboratory verification indicate that IB outbreaks at the end of the egg production period resulted in irreparable decline of production . Marked alterations in exterior and interior quality of the egg have been observed . Both mishapen appearance, rough and thin shell, bleached shell color and watery albumen have occurred . These facts did sometimes cause transient increased "sorting out" of haching eggs . Also fertility rate was noticed to be affected temporarily during and after an attack of IB . In 17 flocks drops of 1.7-22.4% (on an average 7.4%) were registered (fig . 3) . Also hatchability decreased 3.0-43.8% (on an average 13.7%) (fig . 4) . In most of the registered outbreaks of IB the weekly mortality remained below 1%, but in single cases the figure could rise up to about 4% (fig . 5), most often in flocks where complicating factors such as secondary infections caused by E . coli or M . gallisepticum were present. J Bacteriol, 1976 Jul, 127(1), 637 - 43 Association of messenger ribonucleic acid with 70S monosomes from down-shifted Escherichia coli; Jacobson LA et al.; The complexed 70S ribosomes (monosomes) that accumulate in Escherichia coli after an energy source shift-down were examined in an electron microscope . In all cases, the ribosomes lie at or near one end of a ribonucleic acid (RNA) strand . This messenger RNA (mRNA) has a mean length of 168 nm and a length-average length of 200 nm, sufficient to code for polypeptides of a weight-average molecular weight of 20,000 . The length distribution indicates that these strands are a reasonable representation of the population of monocistronic mRNA's of E . coli . The mRNA strands disappear entirely upon digestion with pancreatic ribonuclease, phosphodiesterase I, or polynucleotide phosphorylase . The susceptibility to digestion by 3'-exonucleases indicate that the ribosomes lie at the 5' end of the mRNA strands . These results are consistent with the hypothesis that down-shifted cells have a translational defect at a point subsequent to the binding of ribosomes to mRNA but prior to the formation of the first peptide bond, such that ribosomes remain bound at or near their points of initial attachment to mRNA. J Bacteriol, 1976 Jul, 127(1), 291 - 301 Repression of Escherichia coli carbamoylphosphate synthase: relationships with enzyme synthesis in the arginine and pyrimidine pathways; Pierard A et al.; Cumulative repression of Escherichia coli carbamoylphosphate synthase (CPSase; EC 2.7.2.9) by arginine and pyrimidine was analyzed in relation to control enzyme synthesis in the arginine and pyrimidine pathways . The expression of carA and carB, the adjacent genes that specify the two subunits of the enzyme, was estimated by means of an in vitro complementation assay . The synthesis of each gene product was found to be under repression control . Coordinate expression of the two genes was observed under most conditions investigated . They might thus form an operon . The preparation of strains blocked in the degradation of cytidine and harboring leaky mutations affecting several steps of pyrimidine nucleotide synthesis made it possible to distinguish between the effects of cytidine and uridine compounds in the repression of the pyrimidine pathway enzymes . The data obtained suggest that derivatives of both cytidine and uridine participate in the repression of CPSase . In addition, repression of CPSase by arginine did not appear to occur unless pyrimidines were present at a significant intracellular concentration . This observation, together with our previous report that argR mutations impair the cumulative repression of CPSase, suggests that this control is mediated through the concerted effects of regulatory elements specific for the arginine and pyrimidine pathways. Eur J Biochem, 1976 Jul 1, 66(2), 257 - 68 The use of several energy-coupling reactions in characterizing mutants of Escherichia coli K12 defective in oxidative phosphorylation; Schairer HU et al.; Oxidative phosphorylation, ATP-32Pi exchange, ATP-dependent quenching of acridine-dye fluorescence, ATP-dependent transhydrogenase and ATP-dependent transport of thiomethyl beta-D-galactoside are shown to be experimentally equivalent tools to study the functional state of the ATPase complex in Escherichia coli wild-type and mutant strains defective in oxidative phosphorylation . According to these criteria ten mutants in the ATPase complex were classified having lesions in the unc A,B region of the chromosome . The first mutant type lacks ATPase activity, but the membrane-integrated part of the complex remains functional (class I) . The second mutant type lacks a functional membrane-integrated part, but retains ATPase activity (class II) . The third mutant type is shown to be defective in both parts of the ATPase complex (class III). J Bacteriol, 1976 Jul, 127(1), 162 - 7 Fractionation of membrane vesicles from coliphage M13-infected Escherichia coli; Wickner W; Membrane vesicles were prepared by osmotic lysis of spheroplasts from M13-infected Escherichia coli . Reduced nicotinamide adenine dinucleotide (NADH) oxidase (reduced NAD: oxidoreductase, EC 1.6.99.3) and Mg2+-Ca2+-activated adenosine triphosphatase (ATP phosphohydrolase, EC 3.6.1.3), which are normally localized to the inner surface of the cytoplasmic membrane, were 50% acceesible to their polar substrates in these vesicles . The major coat protein of coliphage M13 is also bound to the cytoplasmic membrane (prior to phage assembly) but with its antigenic sites exposed to the exterior of the cell . Antibody to M13 coat protein was used to fractionate membrane vesicles . Neither agglutinated nor unagglutinated vesicles had altered NADH oxidase and adenosine triphosphatase specific activities . This is inconsistent with such vesicles being a mixture of correctly oriented and completely inverted membrane sacs and suggests that NADH oxidase, adenosine triphosphatase, M13 coat protein, or all three proteins rearrange during vesicle preparation. J Bacteriol, 1976 Jul, 127(1), 555 - 63 Lipoprotein from the outer membrane of Escherichia coli: purification, paracrystallization, and some properties of its free form; Inoyye S et al.; In the envelope of Escherichia coli, is a lipoprotein of molecular weight 7,200 as a major envelope protein . This lipoprotein was previously shown to exist in two different forms in the outer membrane of E . coli: the free form and the boundform, which is covalently linked to the peptidoglycau . The free form of the lipoprotein has been purified and paracrystallized by adding acetone to a sodium dodecyl sulfate solution in the presence of magnesium ion . The paracrystals were needle shaped . An electron micrograph of the negatively stained paracrystals showed a highly ordered ultrastructure . The chemical structure of the free form was compared with that of the bound form by (i) the amino acid composition, (ii) the fatty acid composition, and (iii) the peptide analysis after cyanogen bromide cleavage . The alpha-helical content of the free form of the lipoprotein was measured from the circular dichroism spectrum of the lipoprotein in 0.01% sodium dodecyl sulfate and found to be 87% . Using the purified lipoprotein as antigen, antiserum against the free form of the lipoprotein was obtained . Immunoprecipitation of the lipoprotein with the antiserum was found to be very specific, since only the free form of the lipoprotein was found as a major peak when the antiserum was reacted with the whole envelope proteins solubilized in 0.2% sodium dodecyl sulfate, and the immunoprecipitate thus formed was analyzed by polyacrylamide gel electrophoresis. Mol Biol (Mosk), 1976 Jul-Aug, 10(4), 640 - 6 Interaction of transfer RNA with 50S ribosomal subunits of Escherichia coli in absence of templates; Bresler SE et al.; The equilibrium constant of a complex of tRNA with the 50S ribosomal subunit was measured in the absence of a template . It was shown that the stability of the complex increases with an increase in the concentration of Mg2+, it decreases with an increase in the concentration of univalent ions, and does not depend on the pH of the medium in the range of 7.0-8.2 . Removal of the 3'-terminal nucleoside of tRNA weakens the association approximately 40-fold; the subsequent successive splitting off of another three nucleotides has little effect on the association constant . In 90% 2H2O the stability of the complex increases approximately four-fold, which points to the large contribution of the hydrogen bonds to the free energy of the interaction . The tetranucleotide TphiCG competes slightly with tRNA for sites on the 50S subparticles; this means that the TphiC segment of tRNA does not play an important role in the formation of the complex under investigation. Cell, 1976 Jul, 8(3), 425 - 31 Naturally occurring cross-links in yeast chromosomal DNA; Forte MA et al.; Chromosome-size yeast DNA molecules with a number average molecular weight (Mn) of 3-4 X 10(8) were isolated from sucrose gradients after sedimentation of lysed yeast spheroplasts . Resedimentation showed that the molecules were isolated without introducing appreciable single-strand or double-strand breaks . The presence of cross-links in these molecules was suggested by the observation that the apparent Mn in alkali was greater than expected for separated single strands . Since cross-linked molecules would have strands which fail to separate upon denaturation, this was tested more directly . Neutralization of alkaline denaturing conditions resulted in up to 70% of the intact molecules rapidly reforming duplex structures, as shown by equilibrium banding in CsCI . Experiments with larger E . coli DNA molecules (Mn = 5.2 X 10(8)) indicated that the conditions used were sufficient to denature completely molecules of this size . Results of enzyme treatments suggest that the cross-links are not RNA or protein . Experiments with density-labeled yeast DNA molecules showed that the rapid reformation of duplex DNA is not the consequence either of a bimolecular reaction between separated DNA strands or of intrastrand renaturation . The data indicate that when the yeast DNA molecules are completely denatured, the strands fail to separate . Hence they must be cross-linked . Experiments with sheared DNA show that there are small number of cross-links, one to four, permolecule. Am J Surg, 1976 Jul, 132(1), 64 - 6 Quantitation of local acidosis and hypoxia produced by infection; Raju R et al.; Increased strength of healing incisions infected with E coli was demonstrated in this experiment . Efforts to measure respiratory gas tensions and pH in these incisions were unsuccessful . Therefore, these moieties were measured in normal and infected wound fluid contained in implanted wire cylinders . The wound fluid from infected cylinders was consistently more acidotic and had a lower pO2 and a higher pCO2 than fluid from unifected wound cylinders. Proc Natl Acad Sci U S A, 1976 Jul, 73(7), 2275 - 9 Hydrogen donor system for Escherichia coli ribonucleoside-diphosphate reductase dependent upon glutathione; Holmgren A; E . coli B tsnC 7004, an E . coli B/1 mutant with normal phenotype unable to replicate phage T7 DNA {Chamberlin, M . (1974)J . Virol . 14,509-516}, contained no detectable level of thioredoxin when assayed with ribonucleotide reductase (2'-deoxyribonucleoside-diphosphate:oxidized-thioredoxin 2'-oxidoreductase, EC 1.17.4.1) . Gently lysed E . coli tsnC 7004 cell extracts reduced CDP when supplemented with NADPH as efficiently as the parent strain E . coli B/1 despite the lack of thioredoxin, indicating the presence of another hydrogen transport system . This could be divided into two parts by heat treatment at 85degrees; one heat-stable fraction, which was active in the presence of dithiothreitol or glutathione, and one heat-labile fraction . Addition of yeast glutathione reductase {NAD(P)H:oxidized-glutathione oxidoreductase, EC 1.6.4.2} to the heated extracts restored full activity . The results demonstrate a novel hydrogen transport system in E . coli consisting of NADPH, glutathione, glutathione reductase, and a heat-stable enzyme called "glutaredoxin" . Reduced glutathione at physiological concentrations functions as hydrogen donor for ribonucleotide reduction only in the presence of glutaredoxin . Glutaredoxin was not reduced by E . coli thioredoxin reductase (NADPH:oxidized-thioredoxin oxidoreductase, EC 1.6.4.5) and showed no crossreaction with antibodies against thioredoxin . These results demonstrate the existence of two different electron transfer systems from NADPH to deoxyribonucleotides and provide a function for glutathione in DNA synthesis. Chem Phys Lipids, 1976 Jul, 16(4), 239 - 54 Differential reaction of cell membrane phospholipids and proteins with chemical probes; Marinetti GV et al.; The major aims of this study were to determine the degree of phospholipid asymmetry and the neighbor analysis of phospholipids in different types of cell membranes . For this study a penetrating probe (FDNB), a non-penetrating probe (TNBS) and a cross-linking probe (DFDNB) were used . The reaction of hemoglobin, membrane protein and membrane PE and PS of erythrocytes with DFNB and TNBS was studied over a concentration range of 0.5 to 10 mM probe . TNBS reacts to an extremely small extend with hemoglobin over the concentration range 0.4 to 4 mM whereas FDNB reacts with hemoglobin to a very large extent (50 fold more than TNBS) . The reaction of membrane protein of intact erythrocytes reaches a sharp plateau at 1 mM TNBS whereas the reaction of membrane protein goes to a much larger extent with FDNB with no plateau seen up to 4 mM FDNB . This data shows that TNBS does not significantly penetrate into the cell under our conditions whereas FDNB does penetrate into the cell . The results show that there are four fold more reactive sites on proteins localized on the inner surface of the erythrocyte membrane as compared to the outer surface . TNBS at 0.5 to 2 mM concentration does not label membrane PS and labels membrane PE to a small extent . The reaction of PE with TNBS shows an initial plateau at 2 mM probe and a second slightly higher plateau between 4 to 10 mM probe . TNBS from 0.5-2.0 mM does not react with PS, but between 3 to 10 mM concentration, a very small amount of PS reacts with TNBS . Hence above 2 mM TNBS or FDNB a perturbation occurs in the membrane such that more PE and PS are exposed and react with these probes . These results demonstrate that essentially no PS is localized on the outer surface of the membrane and only 5% of the total membrane PE is localized on the outer surface of the erythrocyte membrane . TNBS and FDNB were reacted with yeast, E . coli, and Acholeplasma cells . With yeast cells, FDNB reacts to a much larger extent with PE than does TNBS, indicating that FDNB penetrates into the cell and labels more PE molecules . With E . coli, but not with erythrocytes or yeast cells, phospholipase A activity was very pronounced at pH 8.5 giving rise to a large amount of DNP-GPE from DNP-PE . A phosphodiesterase was also present which hydrolyized DNP-GPE to DNP-ethanolamine . The multilayered structure of the E . coli cell envelop did not permit a definitive interpretation of the results . It is clear, however, that TNBS and FDNB react to a different extent with PE in this cell . The Acholeplasma membrane had no detectable PE or PS but contains amino acid esters of phosphatidylglycerol . The reaction of these components with TNBS and FDNB indicate that these aminoacyl-PG are localized on both surfaces of the membrane, with 31% being on the outer surface and 69% on the inner surface... J Bacteriol, 1976 Jul, 127(1), 218 - 28 Characterization of group B colicin-resistant mutants of Escherichia coli K-12: colicin resistance and the role of enterochelin; Pugsley AP et al.; Nine classes of group B colicin-resistant mutants were examined to study the role of enterochelin in colicin resistance . Four of the mutants studied (cbt, exbC, exbB, and tonB) hypersecreted enterochelin . Enterochelin hypersecretion was apparently responsible for resistance of the exbC mutant to colicins G and H and for resistance of the exbB mutant to colicins G, H, Ia, Ib, S1, and V . All four mutants scored as colicin B tolerant, even in the absence of enterochelin synthesis . The mutants produced substantially increased amounts of two high-molecular-weight outer membrane polypeptides when grown under limiting iron conditions . The presence of these polypeptides was correlated with increased colicin B-neutralizing activity in the outer membrane preparations. J Bacteriol, 1976 Jul, 127(1), 154 - 61 Adenosine 5'-triphosphate synthesis energized by an artificially imposed membrane potential in membrane vesicles of Escherichia coli; Tsuchiya T et al.; Adenosine 5'-triphosphate (ATP) synthesis driven by an artificially imposed membrane potential in right-side-out membrane vesicles of Escherichia coli was investigated . Membrane vesicles prepared in the presence of adenosine diphosphate were loaded with K+ by incubation with 0.5 M potassium phosphate . Addition of valinomycin resulted in the synthesis of 0.2 to 0.3 nmol of ATP/mg of membrane protein, whereas no synthesis was observed after addition of nigericin . Addition of K+, dicyclohexylcarbodiimide, carbonylcyanide p-trifluoromethoxyphenylhydrazone, or azide to the assay buffer inhibited ATP synthesis . Adenosine diphosphate and Mg2+ were found to be required . Ca2+, which can replace Mg2+ for the hydrolytic activity of the Mg2+-adenosine triphosphatase (ATPase) (EC 3.6.1.3), could not replace Mg2+ in the synthetic reaction and, in fact, inhibited ATP synthesis even in the presence of Mg2+ . Strain NR-70, a mutant lacking the Mg2+-ATPase, was unable to synthesize ATP using an artificially imposed membrane potential . Additionally, the Mg2+-ATPase was found to contain tightly bound ATP. Biochemistry, 1976 Jun 29, 15(13), 2888 - 93 Fluorescence energy-transfer measurements between coenzyme A and flavin adenine dinucleotide binding sites of the Escherichia coli pyruvate dehydrogenase multienzyme complex; Shepherd GB et al.; The interaction of the pyruvate dehydrogenase multienzyme complex from Escherichia coli with 1,N6-etheno-CoA (epsilonCoA) and coenzyme A (CoA) has been investigated using equilibrium binding, steady-state fluorescence, and fluorescence lifetime measurements . A procedure for the resolution of the pyruvate dehydrogenase multienzyme complex into the pyruvate dehydrogenase enzyme and the transacetylase-flavoprotein subcomplex also is given . Direct binding studies with epsilonCoA indicate that 25 bound epsilonCoA molecules/multienzyme complex can be readily displaced by CoA, while approximately 21 bound epsilonCoA molecules/transacetylase-flavoprotein subcomplex can be displaced by CoA . The dissociation constant for the CoA displaceable epsilonCoA is 57.8 muM for the complex and 126 muM for the subcomplex in 0.02 M potassium phosphate (pH 7.0) at 5 degrees C . The kinetic behavior of epsilonCoA as a substrate was investigated and compared with that of CoA under a variety of conditions; the apparent Michaelis constants for epsilonCoA are considerably larger than those for CoA, while the corresponding maximal velocities are smaller . Fluorescence energy transfer measurements between bound epsilonCoA on the dihydrolipoyl transacetylase enzyme and flavin adenine dinucleotide on the dihydrolipoyl dehydrogenase enzyme either in the complex or subcomplex indicate, assuming the emission and absorption dipoles are randomly oriented, that these two probes must be at least 50 A apart. Biochemistry, 1976 Jun 29, 15(13), 2810 - 6 Incorporation of 5'-amino-5'-deoxythymidine5'-phosphate in polynucleotides by use of DNA polymerase I and a phiX174 DNA template; Letsinger FL et al.; An aqueous solution of 5'-amino-5'-deoxythymidine 5'-triphosphate, prepared by incubation of equimolar solutions of 5'-amino-5'-deoxythymidine and sodium trimetaphosphate, stimulates synthesis of acid-precipitable polynucleotides in a system containing single-strand phiX174 DNA template, random oligonucleotide primers, dATP, dCTP,dGTP, Escherichia coli DNA polymerase I, and either magnesium or manganese ion . Approximately onefold synthesis on the template can be achieved and each of the indicated reagents is essential for extensive synthesis . The reaction is slower than the corresponding reaction of dTTP as a consequence of a lower V max and a higher Km for the amino analogue . That aminodeoxythymidine phosphate is incorporated into the synthetic polynucleotides was shown by a double-labeling experiment with {14C}dATP and {32P}-5'-amino-5'-deoxythymidine 5'-triphosphate and by the unusually high lability of the phosphoramidate polynucleotides toward acid . The phosphoramidate polynucleotides range in size from about 100 nucleotide units to well over a thousand nucleotide units, and the size is increased by addition of DNA ligase to the system . These experiments indicate that synthetic polynucleotides in which oligonucleotide blocks have been joined by means of phosphoramidate bonds should prove useful as primers for enzymatic syntheses with DNA polymerase I. Biochemistry, 1976 Jun 29, 15(13), 2804 - 9 The synthesis of 3'-dATP and its use as an inhibitor of ATP-dependent DNA synthesis in toluene-treated Escherichia coli; Gumport RI et al.; A structural analogue of ATP, 3'-deoxyadenosine triphosphate (3'-dATP), has been synthesized from cordycepin (3'-deoxyadenosine), characterized, and determined to be an inhibitor of ATP-dependent DNA synthesis in Escherichia coli cells which have been reduced permeable to nucleoside triphosphates by treatment with toluene . The analogue is a competitive inhibitor of ATP and it inhibits replicative DNA synthesis 50% at concentrations of ca . 0.15 mM in the presence of 1.0 mM ATP and 4 x 10(8) cells/ml . The degree of inhibition of a given amount of 3'-dATP is inversely related to the cell concentration in the reaction mixture . The analogue interferes with some function of ATP which is continuously required during the course of the reaction and does not irreversibly inactivate the cells' DNA synthesis apparatus . 3'-Deoxyadenosine triphosphate may prove useful in elucidating the roles of ATP in DNA synthesis in more purified replicating systems. Biochemistry, 1976 Jun 29, 15(13), 2800 - 3 Comparison of isotope labeling patterns of purines in three specific transfer RNAs; Schoemaker HJ et al.; Purine C-8 tritium-labeling rates have been measured at specific sites in Escherichia coli tRNAIle and tRNA2Tyr . The results are compared with those obtained for yeast tRNAPhe (preceding paper(Gamble et al., 1976)) . The tRNAIle and tRNAPhe fall into the same general class of tRNA structures, while tRNA2Tyr is in a differint class; in particular, the latter is characterized by a large extra loop . In each of the three tRNAs the 3'-terminal A has the same labeling rate and, on a relative basis, appears to be the most rapidly labeled site . Bases in cloverleaf helical sections have markedly retarded labeling rates that collectively fall within an approximately threefold range of time constants . At some of the common purines, believed to be essential for the construction of a general system of tertiary interactions, exchange rates for yeast tRNAPhe are significantly different than those for the two Escherichia coli tRNAs . these differences may arise from variations among the tRNAs in the relative stabilities of specific tertiary interactions, or from other factors as well . In the case of tRNA2Tyr, labeling rates for bases in the large variable region are sufficiently retarded to suggest some structural organization for this part of the molecule . In addition, since exchange rates are similar for some of the bases common to Escherichia coli tRNAIle and tRNA2Tyr, it is likely that the large variable loop of tRNA2Tyr does not interact with or perturb these common sites . Finally, for all three tRNAs, structure formation (e.g., b