Zh Mikrobiol Epidemiol Immunobiol, 1979 Sep, (9), 65 - 8 {Cholesterol synthesis by several strains of Escherichia}; Panchishina MV; The results of the study of lipids in the media used for growing different Escherichia strains are presented . Donor plasma with carbon-labeled sodium acetate was used as culture medium . Those strains which induced an increase in cholesterol content in the medium after 48-hour incubation were considered cholesterin-synthetizing . During the growth of these strains the radioactive marker became incorporated into the lipids accumulated in the medium: phospholipids, cholesterol, fatty acids . The degree of this incorporation depended on the dose of labeled sodium acetate and the amount of the inoculated culture . Cholesterol-synthetizing activity of Escherichia is characteristic of only freshly isolated strains. J Exp Zool, 1979 Sep, 209(3), 367 - 76 The origin of yolk-DNA in Xenopus laevis; Opresko L et al.; Xenopus laevis serum and plasma was found to contain an average of 25 microgram DNA/ml . Isolated X . laevis oocytes incubated in medium containing 25 microgram DNA/ml labeled with either 125I, 32P or 14C and from three different sources (bovine, E . coli and X . laevis), incorporated the label at an average rate of 0.11 ng.mm-2.hr-1 . Sucrose gradient fractionation of oocytes revealed that 40-75% of the acid-precipitable label incorporated was associated with the yolk platelets . Additional incubations of oocytes in unlabeled medium demonstrated that the DNA incorporated into the yolk platelets was undergoing turnover; only 20% of the yolk-associated DNA was still present after a one-week incubation . Our data suggest that yolk-DNA arises by the adventitious uptake of DNA present in the maternal serum by vitellogenic oocytes. J Clin Microbiol, 1979 Sep, 10(3), 275 - 8 Biotyping of Escherichia coli by a simple multiple-inoculation agar plate technique; Buckwold FJ et al.; A nine-test system using multiple-inoculation agar plates for biotyping of Escherichia coli is described . Testing of 959 strains resulted in 78 biotypes . On repeated testing, 96% of 182 strains had identical biotypes or differed by only one test . This system provides satisfactory differentiation among strains and is reproducible . Precise standardization of inoculum size is not required . Multiple inoculation allows time and cost-efficient testing of large numbers of strains. Genetika, 1979 Sep, 15(9), 1578 - 87 {Protective action of colicinogenic factor Ib-P9 on Escherichia coli cells defective in known repair functions after ultraviolet irradiation}; Khmel' IA et al.; The presence of the plasmid colicinogenic factor Ib-P9 in Escherichia coli wild type cells is shown to increase bacterial survival after UV irradiation and the action of N-methyl-N'-nitro-N-nitrosoguanidine . The ability of the plasmid to cause the UV protection is observed in uvrA, uvrB, uvrC, polA, recB, recF E . coli strains, but the plasmid does not restore the UV resistance of the mutant cells to the wild type level . The protective effect of the plasmid CoI Ib-P9 depends on the recA+lexA+ genotype of the cells . The inhibition of protein synthesis (amino acid starvation) before and after UV irradiation does not prevent the UV protection by ColIb-P9 . The nature of the plasmid-associated repair functions is discussed. Am J Med Technol, 1979 Sep, 45(9), 787 - 92 Gastroenteritis due to enteropathogenic, enterotoxigenic, and invasive Escherichia coli: A review; Pickering LK; Escherichia coli that produce diarrhea can be divided into three groups: 1) enteropathogenic, 2) enterotoxigenic, and 3) enteroinvasive . The mechanism of disease production by enteropathogenic E . coli is unknown, but these strains are not presently known to be inherently pathogenic, although they may be important as a cause of gastroenteritis in infants . The two known mechanisms of disease production are elaboration of enterotoxin and mucosal invasion . Heat-labile toxin-producing E . coli are the main cause of diarrhea in travelers while heat-stable toxin-producing E . coli are a cause of scours among new-born swine and cattle . Enteroinvasive E . coli have not been shown to be an important cause of diarrhea in the United States . Enteropathogenic, enterotoxigenic, and enteroinvasive E . coli that currently are associated with diarrhea worldwide may each consist of relatively few serotypes different from those associated with out-breaks of diarrhea in the past . This implies a possible new role for sero-typing of E . coli. J Bacteriol, 1979 Sep, 139(3), 961 - 76 Specificity in formation of type II F' plasmids; Hadley RG et al.; Eight new F' plasmids derived from Hfr strains in which F is integrated at the chromosomal element alpha 3 beta 3 have been isolated and subjected to restriction enzyme, hybridization, and electron microscope heteroduplex analysis . Plasmids carrying extensive amounts of bacterial deoxyribonucleic acid were produced even though they were obtained by selection for transfer of lac, which is closely linked to F in the parental Hfr strains . Seven plasmids were type II Flac+ proC+ purE+ plasmids, and one was a type I Flac+ proC+ plasmid . Five of the Flac+ proC+ purE+ plasmids contain approximately 284 kilobases of bacterial deoxyribonucleic acid, which is identical for all five within the resolution of the restriction enzyme analysis . Theses results indicate that type II F' plasmids are the predominant tra+ F' type from this region of the Escherichia coli K-12 chromosome and that the recombination events leading to formation of these plasmids exhibit site specificity. J Bacteriol, 1979 Sep, 139(3), 850 - 8 Cistrons encoding Escherichia coli heat-labile toxin; Dallas WS et al.; The structure and products of the two cistrons encoding the Escherichia coli heat-labile toxin (LT) were studied . The LT deoxyribonucleic acid (DNA) region had been isolated as part of a DNA fragment from the plasmid P307, and this fragment was joined to the cloning vector pBR313 . Deletion mutations of various lengths were introduced into the LT DNA region and into the adjacent DNA sequences . Analysis of the deletions indicated that the maximum size of the LT DNA region was 1.2 x 10(6) daltons . Two proteins of 11,500 daltons and 25,500 daltons had been shown to be encoded by the LT DNA region . The functions of these LT gene products were investigated . The 11,500-dalton protein had an adsorption activity for Y-1 adrenal cells, and this protein was shown to form aggregates of four or five monomers . The 25,500-dalton protein was shown to have an adenylate cyclase-activating activity . The two cistrons encoding for each of the LT proteins have been located on a genetic map of the LT DNA region . Both cistrons are probably transcribed from the same promoter. J Bacteriol, 1979 Sep, 139(3), 835 - 41 Protein K: a new major outer membrane protein found in encapsulated Escherichia coli; Paakkanen J et al.; The protein composition of purified outer membranes of 47 Escherichia coli strains was examined by sodium dodecyl sulfate-polyacrylamide gradient gel electrophoresis . Of 33 encapsulated strains, all contained an outer membrane protein distinguishable from previously reported proteins . The 14 non-encapsulated strains with one exception lacked this protein . Because of its apparent association with encapsulation (K antigen) we have named it K protein . The protein was purified nearly to homogeneity by chromatography in the presence of detergents, and its composition was determined . Its amino acid composition does not differ significantly from that reported for protein I, another E . coli major outer membrane protein . Furthermore, the N-terminal amino acid sequence of protein K indicates that it is related to protein I. J Bacteriol, 1979 Sep, 139(3), 824 - 34 Properties of Escherichia coli mutants altered in calcium/proton antiport activity; Brey RN et al.; Mutants sensitive to growth inhibition by CaCl2 were found to have alterations in calcium uptake in everted membrane vesicles . These mutations map at different loci on the Escherichia coli chromosomes . A mutation at the calA locus results in vesicles which have two- to threefold higher levels of uptake activity than vesicles from wild-type cells . The calA mutation is phenotypically expressed as increased sensitivity to CaCl2 in a strain also harboring a mutation in the corA locus, which is involved in Mg2+ transport . The calA locus maps very close to purA and cycA at about min 97 . The calB mutation results both in sensitivity to CaCl2 at pH 5.6 and in vesicles with diminished calcium transport capability . The CalB phenotype is also expressed only in a corA genetic background; the calB locus appears to map very near, yet separately from, the calA locus . When the cor+ allele is present, calA and calB mutations still result in a defect in calcium transport in vesicles . In addition, both calC and calD mutations result in vesicles with impaired calcium transport activity . calC is cotransducible with kdp and nagA, whereas calD is cotransducible with proC. J Bacteriol, 1979 Sep, 139(3), 783 - 91 Isolation and characterization of Escherichia coli K-12 mutants unable to induce the adaptive response to simple alkylating agents; Jeggo P; When Esherichia coli cells are exposed to a low level of simple alkylating agents, they induce the adaptive response which renders them more resistant to the killing and the mutagenic effects of the same or other alkylating agents . This paper describes the isolation of one strain that was deficient in mutagenic adaptation and five that were deficient in both mutagenic and killing adaptation, confirming previous suggestions that killing and mutagenic adaptation are, at least to some extent, separable . These six strains have been called Ada mutants . They were more sensitive to the killing and mutagenic effects of N-methy-N'-nitro-N-nitrosoguanidine (MNNG) than the unadapted Ada+ parent . Thus, the adaptation pathway is responsible for circumventing some alkylation-induced damage even in cells that are preinduced . The increase in mutation frequency seen in Ada cells treated with MNNG was the same whether the cells were lexA+ or lexA, showing that the extra mutations found in Ada- strains do not depend upon the SOS pathway . Ada strains accumulated more O6-methyl guanine lesions than the Ada+ parent on prolonged exposure to MNNG, and this supports the idea that O6-methyl guanine is the most important lesion for MNNG-induced mutagenesis . The ada mutations have been shown to map in the 47 to 53-min region of the E . coli chromosome. J Bacteriol, 1979 Sep, 139(3), 1093 - 6 Growth-rate-dependent alteration of 6-phosphogluconate dehydrogenase and glucose 6-phosphate dehydrogenase levels in Escherichia coli K-12; Wolf RE Jr et al.; The levels of 6-phosphogluconate dehydrogenase and glucose 6-phosphate dehydrogenase are subject to metabolic regulation; they increased three- to fivefold with increasing growth rate. J Bacteriol, 1979 Sep, 139(3), 1085 - 8 Interaction between mutant alleles of araC of the Escherichia coli B/r L-arabinose operon; Sheppard DE et al.; Strains were constructed that contain mutational alterations affecting two distinct functional domains within the araC gene protein . The araCi (catabolite repression insensitivity) and araCh (catabolite repression hypersensitivity) mutations were used to alter the catabolite repression sensitivity domain, and mutation to D-fucose resistance was used to alter the inducer binding domain . araCh, D-fucose-resistant double mutants never exhibited constitutive ara operon expression, whereas all of the araCi, D-fucose-resistant double mutants did exhibit constitutivity . When L-arabinose was used as an inducer, most of the double mutants exhibited the sensitivity to catabolite repression associated with the araCi or araCh mutation . However, when D-fucose was used as an inducer, changes in sensitivity to catabolite repression were observed that were attributed to interactions between the two protein domains . The roles of catabolite activator protein and araC gene protein in the induction of the araBAD operon were discussed. J Bacteriol, 1979 Sep, 139(3), 1079 - 81 Effect of transcription on RecBC- and RecF-mediated recombination within the tryptophan operon of Escherichia coli K-12; Cosloy SD; Recombination between tryptophan gene mutations within the trp operon was determined among transductants for an outside linked cysB marker under conditions of repression and derepression . These studies, carried out with recipient strains utilizing the RecBC or RecF pathway, or a combination of these pathways of recombination, demonstrate that transcription of trp genes as regulated by the trp repressor has no significant effect on RecBC- or RecF-mediated recombination within the trp operon. J Bacteriol, 1979 Sep, 139(3), 1049 - 53 Small arrays of electron-dense cylinders in Escherichia coli cells; Bradley DE; Electron microscopy of unstained Escherichia coli cells from cultures kept near 0 degrees C after incubation at 37 degrees C revealed small areas of geometrically arranged electron-dense cylinders . Their morphology, organization, and occurrence are described. Chem Biol Interact, 1979 Sep, 27(1), 1 - 15 Non-random nature of 2-acetylaminofluorene-induced alterations of DNA template capacity; Schwartz EL et al.; Male rats were fed a diet containing 0.03% (w/w) 2-acetylaminofluorene (AAF) and their hepatic DNA was isolated and transcribed with E . coli RNA polymerase . Ingestion of the carcinogen-containing diet for 4 days substantially reduced the template capacity of the isolated DNA . This reduction in template capacity was due to an apparent decreased RNA chain size (up to 50%), with no significant changes in initiation or re-initiation of RNA synthesis . This premature termination of RNA synthesis was accompanied, in some instances, by a reduced rate of RNA chain elongation . When the rats were returned to a basal diet for 7 days following 4 days of AAF ingestion, template capacity and RNA chain size returned to control values . Fractionation of hepatic chromatin on a glycerol gradient revealed that inhibition of DNA template capacity occurs on portions exhibiting characteristics of expressed, as well as those with characteristics of repressed, segments of the genome . In contrast, the DNA isolated from a small, highly condensed chromatin fraction (15% of total chromatin-DNA) showed no significant reduction in total template capacity . Analysis of the fidelity of RNA synthesis on this DNA template was performed by determining the rate of addition of individual nucleotide triphosphates to a growing RNA chain . Large reductions in the rates of adenosine and uridine polymerization were observed while no changes in guanosine or cytidine polymerization were found . This suggests the presence of functionally significant carcinogen-induced modifications of adenine . The inhibition in the rate of adenosine and uridine polymerization was reversed when the animals were placed on a basal diet after AAF ingestion. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4370 - 4 Construction and cloning of rat albumin structural gene sequences; Kioussis D et al.; A recombinant plasmid containing a DNA segment complementary to rat liver albumin mRNA has been constructed, cloned, and used to examine the organization of albumin gene . The 18S fraction of total liver poly(A)-containing RNA was copied into a double-stranded cDNA by avian myeloblastosis virus reverse transcriptase and Escherichia coli DNA polymerase I . The cDNA was inserted into the HindIII site of the plasmid pBR322 via the addition of specific oligonucleotide linkers . Recombinant plasmids were screened by hybrid arrest of mRNA translation and hybridization with specific cDNAs . Thereby, a plasmid was identified that contained a 1200-nucleotide insert corresponding to a segment adjacent to the 5'-terminal region of albumin mRNA . The inserted sequence was used as a hybridization probe to detect five EcoRI fragments of genomic DNA which encode albumin mRNA . These were compared to eight EcoRI fragments identified within the rat genome by albumin cDNA . We conclude that the albumin gene (or genes) is interrupted at more than one site in the coding DNA by intervening sequences . Furthermore, we were able to distinguish those fragments that encode the 5' and 3' ends of the mRNA. Antimicrob Agents Chemother, 1979 Sep, 16(3), 247 - 51 Enhancement of post-ultraviolet killing in Escherichia coli K-12 through the action of gyrase inhibitors: evidence for associated gyrase-recBC deoxyribonuclease function; Purdy MA et al.; This work in conjunction with the results presented in an earlier report (M . A . Purdy and K . L . Yielding, Antimicrob . Agents Chemother . 10:182--184, 1976) showed the following . (i) Nalidixic acid and novobiocin could inhibit post-ultraviolet and post-X-ray survival, implicating gyrase function in deoxyribonucleic acid repair . (ii) The inhibition of post-ultraviolet survival requires the action of functional recBC deoxyribonuclease . (iii) Structural changes in the gyrase could (a) cause recBC mutants to exhibit enhancement of post-ultraviolet killing in the presence of novobiocin, (b) increase the ultraviolet sensitivity of recBC mutants, and (c) enhance the thermal lability of a recBCts mutant. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4308 - 12 A general priming system employing only dnaB protein and primase for DNA replication; Arai K et al.; Priming of phage phi X174 DNA synthesis is effected simply by dnaB protein and primase when the DNA is not coated by single-strand binding protein (SSB) . The five prepriming proteins (n,n',n'',i, and dnaC protein) required for priming a SSB-coated phi X174 DNA circle are dispensable . The dnaB protein-primase priming system is also active on uncoated phage G4 and M13 DNAs and on poly(dT) . Multiple RNA primers, 10--60 nucleotides long, are transcribed with patterns distinctive for each DNA template . Formation of a stable dnaB protein.DNA complex in the presence of primase and ATP supports the hypothesis that dnaB protein provides a mobile replication promoter signal for primase. Mol Biol (Mosk), 1979 Sep-Oct, 13(5), 1021 - 34 {DNA fragmentation of chicken adenovirus CELO by specific endonucleases R . HpaI, R . EcoRI, R . HindIII}; Denisova TS et al.; The effect of specific endonucleases on DNA chiken adenovirus CELO was studed . It was shown that endonucleases R . HpaI, R . EcoRI and R . HindIII cleaved viral DNA into 5,7 and 9 specific fragments, respectively . The sequence of fragments (physical map) was determined and found to be: D-A-E-C-B for enzyme R . HpaI; B-(EG)-C-A-D-F for enzyme R . EcoRI; H-F-A-C-G-B-D-E-I for enzyme R . HindIII. Gene, 1979 Sep, 7(1), 15 - 31 Cloning the argF gene from Escherichia coli K-12 with simian virus 40; Purchio AF et al.; We have inserted a gene coding for ornithine transcarbamylase (OTCase) from Escherichia coli K-12 into the late gene region of simian virus 40 (SV40) DNA and propagated the hybrid molecules as free episomes or by co-infection with an SV40 tsA helper virus . In the first case, the E . coli argF gene was inserted via the EcoRI and BamHI termini in the late gene region of SV40 and the recombinant molecules were used to transfect monkey kidney cells . The hybrid DNA, which was too large to be encapsidated, was replicated for a short time (14 days) but was eventually lost from the surviving cells . In order to allow the argF gene to be packaged into virions, we purified two SV40 vectors containing large deletions of late gene region sequences . One was a 3325 base pair segment from a HaeII + BamHI digest . The argF gene was joined to both vectors at the BamHI site and these linear molecules were used to transfect monkey cells in the presence of SV40 tsA58 DNA as helper . These hybrid DNAs were replicated and packaged into virions . Late in the lytic infection of monkey cells, polyadenylated, cytoplasmic argF transcripts were detected, but significant translation of these trancripts was not observed. Eur J Biochem, 1979 Sep, 99(3), 499 - 505 Regulation of transcription by DNA-bound non-histone nuclear proteins; Crepin M et al.; Purified non-histone proteins from mouse mammary cells bind specifically to homologous DNA or chromatin . Complexes of non-histone protein with DNA or chromatin, isolated on agarose columns, were transcribed with both Escherichia coli RNA polymerase and RNA polymerase B from calf thymus . The fact that complexing of DNA with non-histone proteins increases transcription by E . coli RNA polymerase but not by RNA polymerase B suggests different mechanisms of transcription by these two enzymes . Similar experiments with mouse and Drosophila chromatin indicate that non-histone proteins specifically stimulate the transcription of mouse chromatin by RNA polymerase B . Non-histone proteins stimulate the transcription of mouse mammary tumor virus sequences in chromatin by RNA polymerase B but not by E . coli RNA polymerase . We conclude that those non-histone proteins bound specifically to chromatin are able to activate the transcription of specific genes by eukaryotic RNA polymerase. Arch Environ Health, 1979 Sep-Oct, 34(5), 372 - 6 Effects of silica dust inhalation on the susceptibility of mice to influenza infection; Zarkower A et al.; Mice were exposed to silica dust for durations of up to 36 wk . At intervals of 15, 21, 27, 33, and 36 wk, the ability of the splenic lymphocytes to respond to T and B cell mitogens were determined, and their ability to resist influenza virus infections was determined after 3 and 20 wk of exposure . These exposures had a depressing effect on the response to T cell mitogens at 21 and 27 wk of silica exposure, but there was no effect on the T cell responses after 15, 33, and 36 wk or on B cell responses after all exposure times . No changes could be detected in the ability of the mice to resist pulmonary influenza virus infections or in the survivors ability to form HI antibodies against this virus. J Med Chem, 1979 Sep, 22(9), 1051 - 5 Synthesis and biological properties of N2-substituted spin-labeled analogues of actinomycin D; Sinha BK et al.; We have synthesized N2-{4-(2,2,6,6-tetramethyl-1-piperidinyloxy)}actinomycin D And the related 1,2-diaminoethane and 1,3-diaminopropane derivatives and evaluated their biological properties . Binding studies with the spin-labeled actinomycin D analogues and DNA were carried out by using circular dichroism, electron spin resonance, and thermal denaturation . These studies have suggested that the derivatives bind to DNA and that their DNA-binding modes are similar but not identical . Spin-labeled actinomycin D derivatives were less potent in inhibiting Escherichia coli DNA-dependent RNA polymerase reaction than actinomycin D and were less toxic to L1210 cells in vitro than the parent compound . Spin-labeled actinomycin D derivatives were more common than the parent compounds against P-388 leukemia cells in vitro with little or no toxicity. Z Naturforsch {C}, 1979 Sep-Oct, 34(9-10), 797 - 804 {Quantitative comparison of ribosome binding sites of twelve nucleotide sequences from Escherichia coli (RNA- and DNA phages) based on triplet patterns (author's transl)}; Kohler E; The molecular structure of ribosome binding sites of ten phage genes and two messengers of Escherichia coli were compared concerning the signation parts which are presumably used by ribosomes for recognition and binding . With a simple calculation based on triplet patterns sofar unknown agreements between all of these sequences were found . In several cases it was shown that agreements between old sequences are easier recognizable if the purine- and pyrimidine bases are put into the triplets instead of the four A, G, C, and U (T) bases . In such cases "homologous" parts of sequences were recognized with more distinctness . This is true in our case for the double triplet (hexaplet) py-pu-pu-pu-pu-(pu) and the binding site triplet py-pu-pu, which are preceding the initiator . These triplets are in specific positions in all twelve sequences which were compared . The different course of the quaternary and the binary conformity curves (diagram 1) may show for the investigated area that the RNA phage gene-part is organized according to the well known quaternary triplet code . On the contrary the phage phi-gene-part seems to be organized according to a more simple, binary triplet sequence of purine and pyrimidine bases . The binary sequence seems to be the more original, the quaternary the derived one. Mol Gen Genet, 1979 Sep, 175(2), 203 - 8 Uvm mutants of Escherichia coli K12 deficient in UV mutagenesis . II . Further evidence for a novel function in error-prone repair; Steinborn G; Uvm mutants of Escherichia coli K12 selected for defective UV reversion induction have previously been reported to differ considerably from the UV-reversion-less recA and lexA mutants with regard to survival or mutagenic response to UV, X-rays and alkylating agents . In the present study, the phenotypic characterization of uvm mutants was extended to investigate several cellular processes which also may be related to or involved in UV mutagenesis . Like recA and lexA mutations, the uvm mutations exhibit highly reduced Weigle reactivation and normal host cell reactivation of UV irradiated phage lambda . But unlike recA and lexA, the uvm mutations do not impair genetic recombination, UV induction of prophage lambda or R plasmid-mediated UV resistance and mutagenesis . These phenotypical characteristics and preliminary results of genetic mapping lend further support to the assumption that the uvm site may be a novel locus affecting, apart from the recA and lexA loci, the error-prone repair pathway in E . coli. Mol Gen Genet, 1979 Sep, 175(2), 135 - 44 Characterization of the dnaA, gyrB and other genes in the dnaA region of the Escherichia coli chromosome on specialized transducing phages lambda tna; Hansen FG et al.; Specialized transducing phages lambda tna (tryptophanase) harboring chromosomal DNA and genetic markers from the dnaA region of the Escherichia coli chromosome were isolated . Transductional analysis showed that some of these tnaA transducing phages carry two genes important in DNA replication, namely the dnaA gene (initiation of chromosome replication) and the gyrB gene (subunit B of DNA gyrase), formerly designated couR . The following clockwise order of genetic markers was found: uhp, gyrB, dnaA, rimA, tnaA, bglB . The gene-protein relationship was established by the determination of the gene products encoded on the chromosomal DNA of the different lambda tna . A 54 kD and a 91 kD polypeptide appear to be coded for by the dnaA and gyrB genes, respectively; the 91 kD protein is encoded on a region in which coumermycin sensitivity maps and is with respect to electrophoretic behavior identical to subunit B of DNA gyrase . The 54 kD protein is encoded on the region in which different independently isolated dnaA(Ts) mutations (dnaA5, dnaA46, dnaA167, dnaA203, dnaA204, dnaA205, dnaA211, dnaA508) are located . Additional genes which code for polypeptides with hitherto unknown functions were identified and mapped . The acriflavin sensitivity mutation acrB1 was found to be an allele of the gyrB gene (see "Note Added in Proof"). Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4593 - 7 Sigma subunit of Escherichia coli RNA polymerase affects the function of lambda N gene; Nakamura Y et al.; A new class of Escherichia coli mutants, referred to as grn, has been isolated by localized mutagenesis . These mutations affect the sigma subunit of DNA-dependent RNA polymerase (ribonucleoside 5'-triphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) by abolishing the expression of the lambda N gene, and they are closely lniked to dnaG in the order dnaG-grn-uxaA . Detailed study of one such mutant, grn1, yielded the following results: (i) grn1 is a single mutation and the mutant cell shows cold-sensitivity in growth; (ii) the Grn phenotype of the mutant can easily be suppressed by secondary mutations in the beta subunit gene of RNA polymerase; (iii) purified holoenzyme of RNA polymerase isolated from the mutant showed an altered salt-dependency in vitro, and the mixed reconstitution of the mutant with the wild-type subunits showed that the sigma subunit of the grn1 mutant is altered; (iv) lambda phage mutants (lambda grg), which overcome the grn mutation, can be classified into two groups, the "nin-deletion" and the "N-mutant" groups (both of these are also able to grow on the previously described groN mutant of Georgopoulos and nusAB of Friedman); (iv) the mutant polymerase transcribed 12S as well as 7S RNA from lambda DNA in the presence of the rho factor in vitro . These results indicate that the grn mutation alters the sigma subunit of RNA polymerase and that the sigma subunit participates in activating the N-mediated antitermination mode of lambda phage transcription. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4571 - 5 Transcription promotes recA-independent recombination mediated by DNA-dependent RNA polymerase in Escherichia coli; Ikeda H et al.; The Rpo-mediated recombination of phage lambda takes place independently of the recA function and is promoted by DNA-dependent RNA polymerase of Escherichia coli {Ikeda, H . & Kobayashi, I . (1977) Proc . Natl . Acad Sci . USA 74, 3932--3936} . The crossovers were particularly frequent to the cIII-N and N-cII regions which are transcribed actively . To determine whether the transcription process required for the recombination is the initiation step or the chain elongation step, we have examined the effect of bacterial rho mutation, which affects transcription termination, on the distribution of crossover points in the lambda phage genome . The crossovers in the cII-S interval took place more frequently in rho mutant strains than in wild-type strains . Analysis of lambda mRNA showed that much more O-P-Q mRNA is synthesized in the rho mutant cells than in the wild-type cells and is largely produced by the readthrough from the PR promotor . These results strongly suggest that the chain elongation in transcription plays an essential role in this recombination . Physical analysis of the recombinant phage DNA showed that this recombination is a legitimate type . Models are presented to explain how the transcription complex can promote this recA-independent recombination. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4360 - 4 Cloning of the structural gene (ompA) for an integral outer membrane protein of Escherichia coli K-12; Henning U et al.; The gene (ompA) for the major outer membrane protein II* from Escherichia coli K-12 has been cloned on a 5-megadalton EcoRI fragment by using phage lambda as vector . The gene is expressed during the lytic cycle of the recombinant phage and the insoluble membrane-bound protein was detected in phage plaques with a simple radioimmunoassay . Transfer of the EcoRI fragment into plasmid pSC101 and expression in a host lacking protein II* led to overproduction of protein II* and decreased production of two other major outer membrane proteins . Expression of the plasmid pSC101-ompA+ in minicells derived from an ompA minicell-producing strain led to synthesis, at high rates, of this protein and massive accumulation of a second cell envelope protein most likely representing the biosynthetic precursor of protein II*. Genetika, 1979 Sep, 15(9), 1543 - 54 {New transducing phage with RNA polymerase beta- and beta'-subunit genes derived from a hybrid phage lambda att80: isolation, genetic analysis and physical mapping}; Mindlin SZ et al.; A hybrid lambda att 80 phage with the genetic structure lambda (A-J) phi 80 (att-int-xis) imm lambda..cI857s7 is shown to be a convenient vector for creating transducing phages . On the one hand, the restriction analysis indicates that it has 3 restriction sites for EcoRI in comparison with 5 and 9 sites for parental phages lambda and phi 80 respectively . On the other hand, its buoyant density is less than that of phage lambda and under centrifugation it is easier separated from the phage transducing particles . When lambda att 80 prophage was excluded from the bfe locus of Escherichia coli, transducing phages with genes of two RNA polymerase beta-subunits (rpoB and rpoC) were isolated . To identify the latter, a convenient genetic test was worked out . A physical map of lambda att 80 drifd 35 transducing phage, carrying rpoB and rpoC genes has been constructed using endonucleases EcoRI and HindIII . A comparison of this map and the corresponding maps of transducing phages lambda drifd 18 and lambda drifd 47, studied earlier, led to the discovery of two integration sites of phage lambda within the locus bfe spaced apart by about 1800 nucleotide pairs . At all the sites both phages (lambda and lambda att 80) have integrated in the locus bfe in the counter clockwise order. J Bacteriol, 1979 Sep, 139(3), 1054 - 7 Gene lon and plasmid inheritance in Escherichia coli K-12; Falkinham JO 3rd; Lon- mutants of Escherichia coli K-12 are deficient in the inheritance of F-plasmids by conjugation . This deficiency is distinct from the conjugation deficiency caused by overproduction of capsular polysaccharide which decreases donor-recipient pair formation. Afr J Med Med Sci, 1979 Sep-Dec, 8(3-4), 115 - 23 The risks of umbilical vessel catheterization in a neonatal intensive care unit; Omene JA et al.; Five hundred and fourteen high-risk neonates who had indwelling umbilical catheters at the neonatal intensive care unit of the University of Benin Teaching Hospital were studied . of these 514 neonates, 122 (23.8%) had their catheters in-situ for longer than 24 h . Of the 122, fifty-four (44%) had positive bacterial cultures from their catheter tips . Seven (5.7%) and four (3.2%) of the 122 neonates studied developed septicaemia and necrotizing enterocolitis respectively . Catheterization for periods in excess of 48 h significantly increased the risk of bacterial colonization . Malposition of umbilical catheter tips include: insertion into the right portal vein (thirty-six cases); superior mesenteric vein (three cases) and the left atrium (four cases) . The complications related specifically to the malposition were: air collection in the hepatic venous system (two cases); cardiac arrest (one case); necrotising enterocolitis (one case) and a case of blanching of the abdominal wall . Because of these complications, the indications for catheterization should be restricted to carefully selected patients and strict aseptic technique be adhered to during the procedure. Int J Radiat Biol Relat Stud Phys Chem Med, 1979 Sep, 36(3), 289 - 300 The nature of the damage to Escherichia coli DNA induced by gamma-irradiation; Bresler SE et al.; Quantitative studies of the number of gamma-induced single-strand breaks (SSBs) and enzyme-labile sites (ELSs) were performed using the model of Col E1 plasmids, which undergo transition from the covalently closed form (CCF) into the open circular form (OCF) during gamma-irradiation of the plasmid-bearing strain E . coli JC 411 . By adding 0.5 MEDTA the repair endonucleases of the cell, which effect the transition of ELSs into SSBs during and after gamma-irradiation, were totally inhibited . It was found thless than 15 per cent of the number of gamma-induced lesions are primarily induced SSBs . About the saditions of direct radiation damage . The conclusions are that (1) the contribution of the direct radiation effect in the cell is greater than that of the indirect effect; (2) the main type of gamma-induced lesions are the ELSs (most of which--more than 75 per cent--are alkali-stable; (3) the enzymatic incision of gamma-induced ELSs into SSBs is effected very quickly, mainly during irradiation; and (4) 0.5 MEDTA is a universal inhibitor of repair processes in cell, including the action of N-glycosidases and endonucleases. Carbohydr Res, 1979 Sep, 74, 301 - 7 The ineffectiveness of analogs of D-galactal as competitive inhibitors of, and substrates for, beta-D-galactosidase from Escherichia coli; Dettinger HM et al.; 2,6-Anhydro-3-deoxy-aldehydo-D-lyxo-hept-2-enose (7) and 2,6-anhydro-3-deoxy-D-lyxo-hept-2-enitol (8) were synthesized as half-chair analogs of D-galactal (1) . As 1 is a strong inhibitor of, as well as a substrate for, beta-D-galactosidase from Escherichia coli, the same properties were expected for 7 and 8; however, both were ineffective . This result, together with those of other authors, allows speculative conclusions on the tight binding of 1 to the enzyme only, when water or an alcohol is bound as a co-substrate. J Bacteriol, 1979 Sep, 139(3), 932 - 9 Use of gene fusions to determine a partial signal sequence of alkaline phosphatase; Sarthy A et al.; We have isolated strains of Escherichia coli in which an amino-terminal portion of the cytoplasmic enzyme beta-galactosidase is replaced by an amino-terminal portion of the periplasmic enzyme alkaline phosphatase . The synthesis of these hybrid proteins is regulated by inorganic phosphate and they are located in the cytoplasm . One of these proteins was purified, and 14 amino acids of the amino-terminal sequence were determined . The first five amino acids, Met-Lys-Gln-Ser-Thr, appear to represent a portion of the signal sequence of the precursor of alkaline phosphatase, and the remaining sequence corresponds to that of beta-galactosidase, beginning at amino acid residue 20 . The approach described here could be used for the analysis of signal sequences of exported proteins and for partial amino acid sequence determination of certain of certain other proteins. J Bacteriol, 1979 Sep, 139(3), 721 - 9 Regulation of expression of the flagellin gene (hag) in Escherichia coli K-12: analysis of hag-lac gene fusions; Komeda Y et al.; Previous studies have defined 28 genes necessary for the synthesis of the flagellar apparatus of Escherichia coli K-12 . This study analyzed the influence of the flagellar genes on the expression of the hag gene (structural gene for flagellin) . To this end, a hag::Mu d(Apr lac) mutant which had the lac genes fused to the promoter of the hag gene was constructed . This allowed the measurement of hag gene expression by detection of beta-galactosidase activity . The following observations were made . (i) The hag gene was expressed constitutively in Fla+ cells . (ii) hag gene expression was positively regulated by flaA, FLAB, flaC, flaD, flaE, flaG, flaH, flaI, flaK, flaL, flaM, flaN, flaO, flaP, flaQ, flaR, flaV, flaW, flaX, flaY, flaZ, flbA, and flbB genes.hag-lac expression was not observed in strains with these fla mutations . (iii) The hag gene was expressed in mutants with flaS, flaT, flaU, and flbC defects . Therefore, these genes were not involved in regulation of hag gene transcription. J Bacteriol, 1979 Sep, 139(3), 1014 - 20 Positive control of ilvC expression in Escherichia coli K-12; identification and mapping of regulatory gene ilvY; Watson MD et al.; The construction of a plasmid carrying the ilvC::lacZ fusion is described . This plasmid provides a convenient source of template deoxyribonucleic acid for use in an in vitro protein-synthesizing system . We screened strains deleted in regions of the ilv cluster for their ability to support ilvC-dependent beta-galactosidase synthesis . The fact that two deletions prevented beta-galactosidase production indicated that ilv-C expression is under positive control . By use of plasmids carrying the positive-control factor structural gene ilvY, we were able to restore protein-synthesizing ability to these strains . These plasmids also enabled us to map ilvY between ilvA and ilvC. Clin Chem, 1979 Sep, 25(9), 1649 - 54 Viable and total cell masses in dental plaque as measured by bioluminescence methods; Robrish SA et al.; Bioluminescence methods have been applied to the measurement of the viable and total cell masses of small samples of dental plaque . The total adenine nucleotide content of dental plaque samples and of a pure culture of bacteria was determined and the adenylate energy charge calculated from this . When a pure culture of bacteria was killed with heat, the adenylate energy charge decreased exponentially with duration of treatment and corresponded with a decrease in the count of viable organisms. Tropenmed Parasitol, 1979 Sep, 30(3), 301 - 7 {Immunofluorescent diagnosis of Entamoeba histolytica trophocoites in preserved stool specimens of patients (author's transl)}; Hess U et al.; A fixative and examination technique is described for identification of Entamoeba histolytica trophocoites in faeces with the aid of the indirect immunofluorescence technique . Magnaforms in bloody-dysenteric specimens and Minutaforms in spontaneous or saline purged specimens give equally good fluorescence . The specifity of the rabbit antiserum against axenically grown E . hist . is good: There is strong positive reaction only with trophocoites of E . hist . Weak crossreactions are encountered with trophocoites of E . coli, E . hartmanni and E . polecki . Negative reactions are encountered with trophocoites of Endolimax nana, Jodamoeba bautschlii, and Dientamoeba fragilis . Amebic cysts, flagellates, leucocytes, epithelial cells and Blastocytis give negative reactions, too. Immunology, 1979 Sep, 38(1), 123 - 7 An examination of the O and K specificity involved in the antibody-induced loss of the K88 plasmid from porcine enteropathogenic strains of Escherichia coli; Linggood MA et al.; The heat-labile K88 antigen, a virulence determinant coded for by a transmissible plasmid, was eliminated from enteropathogenic strains of Escherichia coli by passage through media containing antibodies to the heat stable antigens of an Abbotstown (O149:K91,K88ac) strain . The plasmid-curing activity of O149 antisera was not O-antigen specific as O149, O45, O8 and O138 strains of E . coli could be 'cured' of their K88 plasmids by this technique . The curing activity was differentiated from the O-antibody by gel filtration, the O149 antibodies were eluted in the IgM peak while the curing activity was found in the IgG peak . In view of the lack of O-specificity and the absence of K88 antibodies it appears that antibodies to a common heat-stable antigenic determinant were involved in this phenomenon. Mol Biol (Mosk), 1979 Sep-Oct, 13(5), 1070 - 6 {Enzymatic synthesis and characterization of DNA complementary to ceruloplasmin mRNA from rat liver}; Frolova LIu et al.; Poly(A) containing rat liver 21S RNA homogeneous in polyacrylamide gel electrophoresis under denaturing conditions and stimulating the synthesis of ceruloplasmin in a cell-free proteinsynthesizing system, was used as a template for reverse transcription in the presence of T10 primer and highly purified reverse transcriptase from avian myeloblastosis virus . The cDNA made this way was characterized by means of hybridization kinetics with mRNA, by melting of the hybrids formed and by chain length measurements . To increase the degree of representativity, the ceruloplasmin mRNA was fragmented by mild alkaline treatment, enzymatically polyadenylated and transcribed . The cDNA made was fully characterized and the kinetic complexity measured by hybridization with the mRNA was found to be equal to 2300 nucleotides as compared with the value of 3000 nucleotides is expected from gel electrophoresis data . The observed difference may indicate the presence of repeated sequences in the given mRNA . The sufficient representativitness of the synthesized cDNA and its specificity with respect to ceruloplasmin mRNA allows to use it as a molecular probe to study the ceruloplasmin gene structure. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4669 - 73 Requirement for membrane potential in injection of phage T4 DNA; Labedan B et al.; The first stages of infection by phage T4 may be divided into energy-dependent and energy-independent processes . Irreversible adsorption, unplugging, and initial exposure of the DNA terminus may occur at 4 degrees C, or at 37 degrees C in bacteria whose energy-yielding metabolism has been poisoned . DNA injection into the cytoplasm needs higher temperatures and energy from the host cell . The nature of this energy requirements was deduced from the use of metabolic inhibitors . Our results show that T4 DNA injection specifically requires the presence of a protonmotive force across the cytoplasmic membrane of the host . Moreover, the chemical gradient (delta pH) does not appear to be essential, but the membrane potential (delta psi) is required. Proc Natl Acad Sci U S A, 1979 Sep, 76(9), 4544 - 8 A new glnA-linked regulatory gene for glutamine synthetase in Escherichia coli; Pahel G et al.; Mutations in the glnA region of the Escherichia coli chromosome due to Mu prophage insertion result in two phenotypic classes . One class is Gln- and does not synthesize glutamine synthetase{L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2} under any growth condition . The other class produces a low level of glutamine synthetase under all growth conditions and is uncoupled from the regulatory effects of mutations in the glnF and glnD genes . Complementation analysis demonstrates that these two classes of insertions are in different cistrons . From these data we suggest that a regulatory gene, glnG, tightly linked to glnA, mediates both activation and repression of glutamine synthetase synthesis . An analysis of the evidence accumulated to date makes it unlikely that glnG is the only gene in the glnA region involved in the complex system of nitrogen regulation. Science, 1979 Aug 31, 205(4409), 936 - 7 Cytotaxins after the sedimentation behavior of human granulocytes; Catsimpoolas N et al.; Human granulocytes from the peripheral blood of healthy donors were subjected to transient gravity sedimentation analysis in Ficoll density gradient columns (37 degrees C) containing different concentrations of Escherichia coli endotoxin-activated serum and medium 199 . A dramatic serum concentration-dependent dispersion of the cells based on changes in sedimentation velocity was observed as a function of time, using a new optical scanning instrument . The phenomenon was virtually abolished in the presence of cytochalasin B, a known inhibitor of cellular chemotaxis . The width (second statistical moment) of the sedimenting cell distribution increased in a sigmoid fashion as a function of time regardless of cytotaxin concentration . This indicates that a slow and nonlinear response of the granulocytes to the cytotaxins occurs . This new kinetic method should be useful in examining an alternate manifestation of the chemoresponsiveness of phagocytic cells and of cell interactions in general. Biochim Biophys Acta, 1979 Aug 29, 564(1), 31 - 6 The fate of phage lambda DNA in lambda-infected minicells; Witkiewicz H et al.; The fate of phage lambda DNA in lambda-infected Escherichia coli minicells harboring the plasmid ColE1, and in plasmid-free minicells, were studied . Binding of lambda DNA to the minicell membrane, and formation of the supercoiled covalently-closed circular structure has been demonstrated . Phage infection abolishes plasmid DNA synthesis . Only a very slight, non-replicative lambda DNA synthesis occurs, soon after infection . This synthesis is associated with fragments of lambda DNA arising during, or soon after its penetration. Biochim Biophys Acta, 1979 Aug 28, 579(2), 483 - 6 Amino acid sequences of soluble tryptic peptides of chorismate mutase/prephenate dehydratase from Escherichia coli K12; Baldwin GS et al.; The amino acid sequences of 28 soluble tryptic peptides from chorismate mutase/prephenate dehydratase from Escherichia coli K12 have been determined . Together with the four unique cysteine-containing peptides sequenced by Gething and Davidson ((1976) Eur . J . Biochem . 71, 327-336) this accounts for approximately 75% of the total sequence expected for this protein . A high frequency of identify between some of the peptides suggests the possibility of gene duplication during the evolution of the structural gene for the enzyme. Biochim Biophys Acta, 1979 Aug 28, 579(2), 291 - 7 Subunit localizations of zinc(II) in DNA-dependent RNA polymerase from Escherichia coli B; Miller JA et al.; RNA Polymerase holoenzyme and core enzyme from Escherichia coli B have been shown to contain two zinc ions . Flameless atomic absorption spectroscopy of the isolated core subunits indicated that one zinc ion is localized on the beta subunit and the other is bound on the beta' subunit . Atomic fluorescence spectroscopy showed that prolonged dialysis of the metalloenzyme against 0.01 M o-phenanthroline resulted in the removal of both zinc(II) ions with accompanying loss of enzymatic activity . The activity of the apoenzyme was observed to be completely restored by readdition of zinc(II) and partially restored by cobalt(II). Vet Rec, 1979 Aug 25, 105(8), 159 - 64 The pathogenesis of enteric colibacillosis in neonatal unsuckled calves; Pearson GR et al.; The development of pathological lesions in the small intestine of neonatal calves is described . Seven newborn calves were challenged orally with a known enteropathogenic strain of E coli 0101k?(A) and killed at varying times after inoculation . Adhesion of bacteria to the mucosa of the small intestine was observed in all calves . A few organisms were seen in the distal small intestine at three hours after inoculation and thereafter adhesion progressed anteriorly along the intestine in calves killed from six to 36 hours . In these calves pathological changes occurred between six and 12 hours after inoculation . Villi were stunted and thickened and the epithelial surface was irregular . A further calf, anaesthetised from five-and-a-half to 10 hours after inoculation and repeatedly sampled from the distal small intestine, developed similar lesions abruptly at nine hours after inoculation . Villus and crypt lengths in the challenged calves were compared with those in three normal uninoculated control calves. J Biol Chem, 1979 Aug 25, 254(16), 8074 - 82 Purification and properties of L-Aspartate-alpha-decarboxylase, an enzyme that catalyzes the formation of beta-alanine in Escherichia coli; Williamson JM et al.; L-Aspartate-alpha-decarboxylase, an enzyme that catalyzes the production of beta-alanine, has been purified to apparent homogeneity from Escherichia coli . The properties of the enzyme are: (a) pH optimum of 6.8 to 7.5, (b) temperature optimum of 55 degrees C, (c) Km for L-aspartate of 0.16 mM, and (d) molecular weight of 58,000 . The activity of the enzyme is inhibited by reagents (hydroxylamine, phenylhydrazine, and sodium borohydride) that react with carbonyl groups, but no pyridoxal phosphate is present . The compound containing the carbonyl group has been identified as covalently bound pyruvate . Approximately 1 mol of pyruvate was found/mol of enzyme . That the enzyme has a biosynthetic function rather than a catabolic role is indicated by the observations that a mutant (designated as E . coli 99-2) which requires either beta-alanine or pantothenic acid for growth contains only trace amounts of enzyme activity, whereas it is present in substantial amounts in the parent strain (E . coli W) and in a spontaneous revertant of the mutant. J Biol Chem, 1979 Aug 25, 254(16), 7820 - 6 The effect of template secondary structure on vaccinia DNA polymerase; Challberg MD et al.; Vaccinia virus DNA polymerase will utilize a substrate consisting of phi X174 DNA primed with a strand of a unique restriction fragment, but the reaction is inefficient . Examination of the reaction products by alkaline agarose gel electrophoresis revealed a few discrete fragments, each corresponding to an extended primer strand . This result implies that specific barriers exist on the phi X174 template which impede, but do not completely halt, the progress of the enzyme . Only a few per cent of the template molecules were completely copied . Similar findings were reported by Sherman and Gefter using Escherichia coli DNA polymerase II and fd DNA (J . Mol . Biol . (1976) 103, 61-76) . Several observations suggest that the barriers are regions of template secondary structure . Some barriers are more effective than others, and they increase in both effectiveness and number as the temperature is decreased . The same barriers are observed with T4 DNA polymerase, but none are detected with E . coli DNA polymerase I . Finally, the major barriers are located in regions of the phi X174 sequence known to contain hairpin structures of relatively high stability . The exact stopping point of one of the major barriers is within the duplex stem of a hairpin structure . These results show that DNA polymerases are a useful probe of the secondary structure of a single-stranded DNA. J Biol Chem, 1979 Aug 25, 254(16), 7752 - 7 Conformational and ligand binding properties of the isolated domains from the beta 2 subunit of Escherichia coli tryptophan synthetase investigated by the reactivity of their cysteines; Goldberg ME et al.; A mild proteolytic treatment of the dimeric beta 2 subunit of Escherichia coli tryptophan synthetase (L-serine hydrolase (adding indole) EC 4.2.1.20) is known to nick each polypeptide chain into two complementary fragments, F1 and F2 (Hogberg-Railbaud, A., and Goldberg, M.E . (1977) Proc . Natl . Acad . Sci . U.S.A . 74, 442-446) . The reactivity of the cysteines in the isolated or associated fragments is studied and used to characterize the structural and functional properties of these fragments . It is shown that the total number of cysteines, their reactivity to dithiobisnitrobenzoate, and their protection by various ligands are the same in the nicked and intact enzyme, thus demonstrating the close structural analogy between these two proteins . In the isolated F1 fragments two cysteines are reactive and two are buried, thus confirming that this fragments has a compact, globular structure . Various ligands tested fail to produce any modification of the cysteines in the isolated fragments, thus suggesting that none of the fragments alone carries a binding site for the substrates and coenzyme. J Biol Chem, 1979 Aug 25, 254(16), 7529 - 33 Location of the sugar-binding site of L-arabinose-binding protein . Sugar derivative syntheses, sugar binding specificity, and difference Fourier analyses; Newcomer ME et al.; The sugar-binding site of the L-arabinose-binding protein, an essential component of the high affinity L-arabinose uptake system in Escherchia coli, is located deep in a cleft formed by the asymmetric contributions from both of the two similar domains . The site was unambiguously identified with the electron-rich substrate analog 6-bromo-6-deoxy-D-galactose in a difference Fourier analysis . The observation that the original native structure might have been solved with bound L-arabinose necessitated the synthesis of a heavy atom analog, its structure consistent with the known sugar-binding specificity of the protein . Difference Fourier maps (3.5 A) of crystals soaked in 46 mM analog showed a peak 3.5 times background, which is attributed to the -CH2Br moiety of the analog . Superposition of a difference map onto a 2.8-A native electron density map indicated that the difference peak is 6 to 7 A from the reactive single cysteine (Cys-64) and partially coincident with an "extraneous" density found in the native map . This "extraneous" peak was previously attributed to a bound L-arabinose molecule, and its presence accounts for the early failures of difference Fourier analyses of crystals soaked in or co-crystallized with L-arabinose to locate the sugar-binding site. J Biol Chem, 1979 Aug 25, 254(16), 7485 - 7 Differential inhibition of host and viral thymidylate synthetases by folylpolyglutamates; Maley GF et al.; The ability of folate analogues to inhibit host and viral thymidylate synthetases was measured using the corresponding Escherichia coli and T2-phage-induced enzymes . In the absence of Mg2+, 6 x 10(-7) M pteroylhexaglutamate inhibited the T2-phage-induced synthetase by 50%, but at least 100-fold greater levels of this compound were necessary to inhibit the E . coli synthetase by this amount . At 2.5 x 10(-6) M pteroylhexaglutamate, at least 80% inhibition of the T2-phage synthetase could be obtained with little or no inhibition of the E . coli enzyme . The pteroylmonoglutamate was about 2 orders of magnitude less inhibitory towards the T2-phage enzyme than the pteroyltri- to -heptaglutamates . However, upon addition of Mg2+ to the assay mixture, the inhibition produced by pteroylhexaglutamate was essentially reversed, with the E . coli synthetase now increasingly inhibited by this compound and the T2-synthetase only minimally impaired . Methotrexate and N10-formyl-2-amino-4-hydroxyquinazoline, although inhibitory to both enzymes in the presence or absence of Mg2+, did not show this differential selectivity . These results suggest that certain folate analogues may be useful in distinguishing between a host and an infecting organism's thymidylate synthetase and could thus provide an additional means of screening for potential chemotherapeutic agents. J Biol Chem, 1979 Aug 25, 254(16), 7465 - 7 A simple and rapid purification method for Escherichia coli DNA polymerase I; Rhodes G et al.; We report a simple, three-step method for the purification of Escherichia coli DNA polymerase I . Its advantages over other procedures are ease and rapidity, the absence of an autolysis or any high speed centrifugation step, and applicability to large quantities of material . In addition, RNA polymerase can be isolated as a by-product . We have applied this method to purify DNA polymerase both from wild type E . coli cells and from cells bearing a lambda prophage carrying the polA gene (Kelley, W.S., Chalmers, K., and Murray, N.E . (1977) Proc . Natl . Acad . Sci . U.S.A . 74, 5632-5636) . This latter source amplifies the amount of DNA polymerase in the cells by at least 10-fold. J Biol Chem, 1979 Aug 25, 254(16), 7915 - 20 Purification and properties of phosphorylated isocitrate dehydrogenase of Escherichia coli; Garnak M et al.; Phosphorylated NADP+-isocitrate dehydrogenase (EC 1.1.1.42) has been purified to electrophoretic homogeneity from in vivo 32P-labeled Escherichia coli . The cells used as the source of phosphorylated enzyme were harvested 1 h after the addition of 5 mCi of {32P}orthophosphoric acid and 25 mM sodium acetate to cultures grown to early stationary phase on a low phosphate medium with limiting glucose . Double immunodiffusion and autoradiography demonstrated immunological identity between the 32P-labeled NADP+-isocitrate dehydrogenase and the enzyme isolated from glucose-grown E . coli . The phosphoenzyme had an apparent subunit molecular weight of 51,000 as determined by denaturing acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, and the radioactivity co-electrophoresed with NADP+-isocitrate dehydrogenase activity when purified enzyme was subjected to nondenaturing gel electrophoresis . {32P}Phosphoserine was identified following partial acid hydrolysis of the purified phosphoenzyme. Nucleic Acids Res, 1979 Aug 24, 6(12), 3759 - 84 Cloning of chicken embryo tRNA genes using single stranded nucleosomal DNA highly enriched for tRNA complementary sequences; Wittig B et al.; DNA from chicken embryo nucleosome tetramers (about 760 base pairs in size) was enriched for tRNA genes by RPC-5 chromatography . The enriched DNA was hybridized with chicken embryo total tRNA and the hybridized DNA isolated utilizing a) avidinbiotin interaction, b) diazobenzyloxymethyl paper, and c) high temperature RPC-5 chromatography . The obtained single stranded DNA highly enriched for tRNA complementary sequences was hybridized with total DNA from nucleosome monomers (140--190 base pairs in size) and the excess of non hybridized monomer nucleosome DNA removed by Sepharose 4B chromatography . The hybrid molecules obtained were made fully double stranded by incubation with E . coli DNA polymerase I, DNA ligase, and exonuclease III . DNA was inserted into plasmid pBR322 by G-C joining procedure and the recombinant DNA used to transform the E . coli strain chi 1776 . More than 70% of the transformants obtained hybridize to chicken embryo total tRNA. Biochemistry, 1979 Aug 21, 18(17), 3714 - 23 Mechanism of phenylalanyl-tRNA synthetase of Escherichia coli K . 10 . Modulation of catalytic properties by magnesium; Pimmer J et al.; The association of phenylalanylptRNA and Mg2+ follows a biphasic concentration dependence as indicated by the active site directed fluorescent indicator 2-p-toluidinyl-naphthalene-6-sulfonate . The macroscopic dissociation constants are 0.16 +/- 0.03 and 4.1 +/- mM . The effect of Mg2+ on the association of enzyme and MgATP, on the synergistic binding of MgATP and L-phenylalaninol, and on the pre-steady-state synthesis and pyrophosphorolysis of the enzyme-phenylalanyladenylate complex in the absence and the presence of tRNA Phe has been measured by established equilibrium and stopped-flow techniques using 2-p-toluidinylnaphthalene-6-sulfonate . At 10 mM Mg2+, the association of enzyme and MgATP is biphasic with dissociation constants of 0.25 +/- 0.03 and 9.1 +/- 1.7 mM . At 2 mM Mg2+, a single dissociation constant of 5.0 +/- 0.5 mM is indicated . The coupling constant of the synergistic reaction is 15 at 1 mM Mg2+ and 290 at 10 mM Mg2+ . The Hill constant of the sigmoidal dependence is 3.6 . The strengthening of the synergism is believed to reflect a Mg2+-dependent coupling of the synergistic reactions at the two active sites of the enzyme, the coupling being negligible at 1 mM and maximal at 10 mM Mg2+ . The pre-steady-state rate of adenylate synthesis is accelerated by the presence of Mg2+ . The effect is to decrease the value of the Michaelis-Menten constant of MgATP . Another effect is to increase the rate constant when tRNA Phe is present . At subsaturating {MgATP}, the {Mg2+} dependence of the observed rate constant is hyperbolical in the absence and sigmoidal (Hill constant, 3.5) in the presence of tRNA Phe . The rate of the pyrophosphorolysis is enhanced by a decrease of the Michaelis-Menten constant of MgPPi . The effects on the thermodynamics and kinetics parallel the occupancy of the low-affinity Mg2+-binding sites of the enzyme. Biochem J, 1979 Aug 15, 182(2), 493 - 502 Abnormal ribosome assembly in a mutant of Escherichia coli; Butler PD et al.; The mutant strain, 15--28, of Escherichia coli accumulates ribonucleoprotein ('47S') particles that were previously shown {Markey, Sims & Wild (1976) Biochem . J . 158, 451--456} to be an unusual intermediate in the assembly of 50S ribosomal subunits... Biochem J, 1979 Aug 15, 182(2), 407 - 12 Biosynthesis and turnover of outer-membrane proteins in Escherichia coli ML308-225; Allen RJ et al.; Isolated outer membranes and outer-membrane extracts from Escherichia coli ML308-225 in the early-exponential growth phase contain more protein than do corresponding preparations from late-exponential- or stationary-phase bacteria . Isotope-dilution experiments show that this is due to a loss of protein from the membrane during the exponential growth phase . Inhibition of bacterial growth and protein synthesis stabilizes the outer-membrane-protein concentration . Protein synthesis in the absence of bacterial growth results in higher concentrations of protein in the outer membrane. J Am Vet Med Assoc, 1979 Aug 15, 175(4), 388 - 91 Peritoneal lavage in the horse; Valdez H et al.; Eight horses ranging in age from 4 days to 9 years were treated for peritonitis . Escherichia coli was isolated in four cases and Nocardia sp in one case . In each case, a catheter placed in the peritoneal cavity allowed drainage of a large amount of purulent fluid . Retrograde peritoneal lavage was performed through a Foley catheter or medical tubing, using Ringer's lactate solution containing kanamycin, povidone iodine, or nitrofurazone . All except two horses responded well to repeated lavage. Eur J Biochem, 1979 Aug 15, 99(1), 39 - 47 Iron supply of Escherichia coli with polymer-bound ferricrocin; Coulton JW et al.; Uptake of ferric iron from ferricrocin was studied in Escherichia coli using a polymer-coupled ferricrocin that was unable to penetrate into the cell . Ferricrocinyl polyethylene glycol succinate (Mr 7000 -- 8500) promoted growth of E . coli K-12 AB2847 aroB under iron-limiting conditions . In iron-starved cells, uptake of 55Fe could be demonstrated; the amount of iron accumulated amounted to 10% of that observed with free ferricrocin . The iron supply by ferricrocin bound to polyethylene glycol was strictly dependent upon the functions expressed by the tonA and the tonB genes, as was the iron uptake promoted by free ferricrocin . Polymer-bound ferricrocin protected cells against colicin M and phage T5 by competition for the common tonA-coded outer membrane receptor protein . In addition, the rate of iron transport via the negatively charged ferricrocinyl succinate was as fast as via the neutral ferricrocin molecule . No ligand was found associated with the cells . Penetration of chelator beyond receptor is not necessary for siderophore-mediated iron uptake . It is concluded that sufficient amounts of iron can be released from the polymer complex to satisfy growth requirements. Eur J Biochem, 1979 Aug 15, 99(1), 187 - 201 On the binding of tRNA to Escherichia coli RNA polymerase; Spassky A et al.; The fixation of tRNA to Escherichia coli RNA polymerase has been investigated . Bound and free tRNA have been separated and quantified after filtration through cellulose nitrate filters, centrifugation or sucrose gradients or electrophoresis in polyacrylamide gels . We detect no differences between the fixation of E . coli fMet-tRNAfMet, Met-tRNAmMet or uncharged unfractionated tRNA to RNA polymerase . Tight complexes, with a long residence time, are formed between core enzyme and tRNA with a dissociation constant of less than 1 nM . Complexes exist between tRNA and both monomer and dimer forms of the core enzyme . In the monomer complex, one tRNA is bound per alpha 2 beta beta' unit, whereas in the dimer complex only 0.5 tRNA molecule is fixed per alpha 2 beta beta' unit . In contrast to the core enzyme, very little tRNA fixes tightly to the holoenzyme at salt concentrations greater than 80 mM . At lower salt concentrations tRNA fixation results in a loss of sigma subunit from the holo enzyme to the resulting core enzyme where it binds tightly . DNA fixation reduces the binding of tRNA to RNA polymerase and tRNA fixation reduces the binding of DNA . However, binding of DNA to polymerase is not competitive with binding of tRNA, and ternary complexes between RNA polymerase, DNA and tRNA are shown to exist . Our results are discussed in relation to other studies concerning the effects of tRNA upon RNA polymerase. Eur J Biochem, 1979 Aug 15, 99(1), 105 - 11 Isolation of plasmid-protein complexes from Escherichia coli; Busby S et al.; A procedure is described for the isolation of complexes between pMB9 plasmids and protein from Escherichia coli which are stable during centrifugation on sucrose gradients and are not destroyed in the presence of competitor DNA . The proteins in these complexes have been analysed by dodecyl sulphate/polyacrylamide gel electrophoresis . Only 10 polypeptide species are found in significant quantities, many of which are bound to both the plasmid and host DNA . We have also detected the presence of one protein which binds to a specific DNA sequence inserted in the plasmid. Biochim Biophys Acta, 1979 Aug 14, 547(2), 198 - 210 Localization and characterization of cytochromes from membrane vesicles of Escherichia coli K-12 grown in anaerobiosis with nitrate; Sanchez Crispin JA et al.; Cytochromes b of anaerobically nitrate-grown Escherichia coli cells are analysed . Ascorbate phenazine methosulfate distinguishes low and high potential cytochromes b . Reduction kinetics performed at 559 nm presents a very complex pattern which can be analysed assuming that at least four b-type cytochromes are present . The electron transport chain from formate to oxygen would contain a low potential cytochrome b-556, a cytochrome b-558 associated to the oxidase, and a cytochrome d as the principle oxidase . Cytochrome o is also present, but seems to be functional only at low oxygen concentrations . A cytochrome b-556 associated to nitrate reductase is shown to belong to a branch of the formate-oxidase chain . 2-N-Heptyl-4-hydroxyquinoline-N-oxide affects the reduction kinetics in a very complex way . One inhibition site is in evidence between cytochrome b-558 and cytochrome d; another between the cytochrome associated to nitrate reductase and the nitrate reductase . A third inhibition site is located in the common part of the formate-nitrate and the formate-oxidase systems . Ascorbate phenazine methosulfate is shown to donate electrons near cytochrome b-558. J Chromatogr, 1979 Aug 11, 176(2), 217 - 24 Rapid assay for tryptophanase using reversed-phase high-performance liquid chromatography; Krstulovic AM et al.; A rapid and sensitive high-performance liquid-chromatographic assay for tryptophanase based upon the fluorometric measurement of the enzymatically liberated indole was developed . The total incubation time is 20 min, and the reversed-phase separation is fast (elution time of indole in 8 min) and reproducible . The sensitivity of the method is in the nanomole range . This method was tested in the assay of tryptophanase activity in E . coli, giving an average activity of 6589.6 U/g of cells . Because of its speed, high sensitivity and minimal sample preparation, this method circumvents several problems commonly encountered in standard spectrophotometric methods of analysis. Nucleic Acids Res, 1979 Aug 10, 6(11), 3673 - 84 Release of 7-methylguanine residues whose imidazole rings have been opened from damaged DNA by a DNA glycosylase from Escherichia coli; Chetsanga CJ et al.; Double-stranded DNA containing 7-methylguanine residues whose imidazole rings have been opened, i.e . 2,6-diamino-4-hydroxy-5-N-methylformamido-pyrimidine residues, may be prepared by treatment of DNA with dimethyl sulfate followed by prolonged incubation at pH 11.4 . These substituted formamidopyrimidine residues are actively removed from DNA by a DNA glycosylase present in E . coli cell extracts . The enz;me shows no apparent cofactor requirement and has a molecular weight of about 30 000 . The release of ring-opened 7-methyl-guanine residues is due to a previously unrecognized activity, different from the three known E . coli DNA glycosylases that release uracil, 3-methyladenine, and hypoxanthine from DNA . This enzyme may serve to repair a major secondary alkylation product in DNA . In addition, it may remove nonmethylated purines, whose imidazole rings have been opened, from X-irradiated DNA. Nucleic Acids Res, 1979 Aug 10, 6(11), 3641 - 50 Binding of Escherichia coli ribosomal proteins to 23S RNA under reconstitution conditions for the 50S subunit; Marquardt O et al.; The RNA binding capacity of 50S proteins from E . coli ribosomes has been tested under improved conditions; purified proteins active in reconstitution assays were used, and the binding was studied under the conditions of the total reconstitution procedure for the 50S subunit . The results are: 1) Interaction of 23S RNA was found with 17 proteins, namely L1, L2, L3, L4, L7/L12, L9, L10, L11, L15, L16, L17, L18, L20, L22, L23, L24 and L29 . 2) The proteins L1, L2, L3, L4, L9, L23 and L24 bound to 23S RNA at a level of about one copy per RNA molecule, whereas L20 could bind more than one copy (no saturation was observed at 1.8 copies per 23S RNA), and the other proteins bound 0.2--0.6 copies per RNA . 3) L1, L3, L7/L12 showed a slight binding to 16S RNA, L26 (identical with S20) strong binding to 16S RNA . 4) The binding of L2, L7/L12, L10, L11, L15, L16 and L18 was preparation sensitive, i.e . the binding ability changed notably from preparation to preparation . 5) All proteins bound equally well to 23S RNA in presence of 4 and 20 mM Mg2+, respectively, except L2, L3, L4, L7/L12, L9, L10, L15, L16 and L18, which bound less strongly at 20 mM than at 4 mM Mg2+. Nucleic Acids Res, 1979 Aug 10, 6(11), 3459 - 69 Nucleotide sequence of starfish initiator tRNA; Kuchino Y et al.; The nucleotide sequence of starfish ovary initiator tRNA was determined to be pA-G-C-A-G-A-G-U-m1G-m2G-C-G-C-A-G-U-G-G-A-A-G-C-G-U-G-C-U-G-G-G-C-C-C-A-U-t6A-A-C-C-C-A-G-A-G-m7G-D-m5C-C-G-A-G-G-A-psi-C-G-m1A-A-A-C-C-U-C-G-C-U-C-U-G-C-U-A-C-C-AOH . The sequence was determined by a combination of the two different post-labeling techniques . Two-dimensional cellulose thin-layer chromatography was adopted for analysis of 5'-terminal nucleotides of tRNA fragments produced by formamide treatment . The nucleotide sequence of starfish initiator tRNA is very similar to that of mammalian cytoplasmic initiator tRNAs, but has seven different nucleotide residues and two modifications: residue 55 is psi instead of U, and residue 26 is unmodified G instead of m2G. Nucleic Acids Res, 1979 Aug 10, 6(11), 3443 - 58 Rapid print-readout technique for sequencing of RNA's containing modified nucleotides; Gupta RC et al.; A rapid, simple, and highly sensitive method for sequence analysis of RNA was developed, which consists of the following steps: (i) controlled hydrolysis of the RNA by brief heating in water; (ii) (32P)-labeling of 5'-hydroxyl groups of the fragments produced in (i); (iii) resolution of labeled fragments by size on polyacrylamide gels giving the familiar "ladder"; (iv) contact transfer ("print") of the ladder from the gel to a PEI-cellulose thin layer; (v) in situ treatment of the ladder with RNase T2 resulting in the release of 5'-(32P)-labeled nucleoside-3',5' diphosphates; (vi) contact transfer and thin-layer separation of (32P)-labeled nucleotides on PEI-cellulose in ammonium sulfate and ammonium formate solvents; (vii) autoradiography . The chromatographic behavior of the 4 major and 18 modified nucleotides was determined . The positions of major and modified nucleotides in the sequence can be read directly from the separation patterns displayed on X-ray film . As this is the only sequencing method presently available that allows one to display and identify directly the positions in the RNA chain of major and modified nucleotides, no additional procedures are required to analyze the latter. J Biol Chem, 1979 Aug 10, 254(15), 7377 - 87 Role of a membranous sialyltransferase complex in the synthesis of surface polymers containing polysialic acid in Escherichia coli . Temperature-induced alteration in the assembly process; Troy FA et al.; Membrane-associated sialyltransferase complexes of Escherichia coli K-235 catalyze the synthesis of sialyl polymers which remain associated with the cell envelope . Sialyl monophosphorylundecaprenol is an intermediate in the formation of these unique surface structures, and fluidity of the lipid phase is required for the proper function of the enzyme complex (Troy, F.A., Vijay, I.K., and Tesche, N . (1975) J . Biol . Chem . 250, 156-163, 164-170) . In membranes containing an increased unsaturated fatty acid content of the phospholipids, obtained by growing cells at 15 degrees C, synthesis of polysialic acid was uncoupled from synthesis of the sialyl lipid-linked intermediate . Using reconstruction experiments, the importance of the role of an endogenous acceptor in polymer formation was suggested by the unexpected finding that polysialic acid synthesis could be reactivated in inactive membranes by the addition of an exogenous acceptor which contained sialic acid . Concomitant with polymer synthesis was a rapid loss of labeled sialic acid from the lipid phase . The activated sialic acid was shown to be transferred directly to the exogenous acceptor . These results establish: 1) that the temperature-induced alteration in polymer synthesis resulted from the inability of cells grown at 15 degrees C to either synthesize or assemble a functional endogenous acceptor and not from a defect in the synthesis of the sialyltransferase; 2) the intermediate precursor role of lipid-soluble sialic acid in sialyl polymer synthesis; and 3) that the exogenous acceptor served directly as an "acceptor" and not as a catalytic "effector" which stimulated an inactive membrane-enzyme complex . These results are in accord with the possibility that the low temperature-induced derangement in polymer formation is a consequence of the altered lipid structure resulting from the greater unsaturated fatty acid content in the membrane phospholipids . U-14C-labeled exogenous acceptor was isolated from the culture filtrate of cells grown at 37 degrees C and purified to homogeneity by preparative polyacrylamide gel electrophoresis . The pure acceptor was characterized structurally as a homopolymer of sialic acid with a degree of polymerization of approximately 12 . Potassium borohydride reduction of the acceptor prior to complete hydrolysis with neuraminidase established that the polymer possessed a free reducing terminus of sialic acid . Subsequent structural studies showed that these oligomers of sialic acid appeared in the culture filtrate as a result of acid-catalyzed hydrolysis from membrane-associated polysialic acids of about 150 to 200 sialyl residues . Marked diminution of several membrane proteins was observed for cells grown at 15 degrees C . The possible relationship of these alterations to the upward shift in unsaturated lipids and to the loss of a functional endogenous acceptor is currently under study. J Biol Chem, 1979 Aug 10, 254(15), 7123 - 8 Preparative enzymatic synthesis and hydrophobic chromatography of acyl-acyl carrier protein; Rock CO et al.; We have used purified preparations of acyl-acyl carrier protein synthetase to prepare pure, native acyl-acyl carrier proteins (acyl-ACP) ranging in chain lengths from C10:0 to C delta 9 18:1 . Factors affecting yield are explored and reaction conditions are presented that yield 0.8 to 0.9 mg of C16:0-ACP/ml of reaction mix . Ohter acyl groups, such as C10:0 and C delta 9 18:1 are poorer substrates and gave correspondingly lower yields . Acyl-Acp synthetase may be recovered from the reaction mixture using blue-Sepharose CL-6B and recycled . ACP and acyl-ACP are separated by hydrophobic chromatography on octyl-Sepharose CL-4B . Mixtures of acyl-ACPs could be resolved according to acyl chain length using octyl-Sepharose CL-4B columns eluted with a 2-propanol gradient . The high resolution obtained using 2-propanol gradients to separate acyl-ACP species suggests that similar techniques would be applicable to the chromatography of protein mixtures on hydrophobic supports. J Biol Chem, 1979 Aug 10, 254(15), 7111 - 5 The effect of chemical modification of 3-(3-amino-3-carboxypropyl)uridine on tRNA function; Friedman S; The minor base 3-(3-amino-3-carboxypropyl)uridine (acp3U) in Escherichia coli tRNAPhe was acylated with the N-hydroxysuccinimide esters of acetic, phenoxy-acetic, and naphthoxyacetic acid, as well as the ester of 5-dimethylaminonaphthalene-1-sulfonyl (dansyl)-glycine . The derivatives of tRNAPhe formed were all capable of accepting phenylalanine . There were only minor effects on the kinetic parameters of these derivatives for E . coli phenylalanyl-tRNA synthetase . There was no effect on the ability of tRNAPhe to participate in poly(U)- or poly(ACU)-directed polypeptide synthesis or in the poly(U)-stimulated binding to E . coli ribosomes . The rate of photodynamic cross-linking of 4-Srd 8 to Cyd 13 was decreased in tRNAs containing the acetyl and dansyl-glycyl derivatives of acp3U, indicating that acylation of this base may perturb the tertiary structure of the tRNA . This base in tRNAPhe does not appear to play any role in the known biological functions of tRNAPhe. J Biol Chem, 1979 Aug 10, 254(15), 6894 - 903 Both positional isomers of aminoacyl-tRNA's are bound by elongation factor Tu; Alford BL et al.; Six purified Escherichia coli and yeast tRNA's were converted to positionally defined tRNA's terminating in 2'- and 3'-deoxyadenosine; the modified (amino-acyl) tRNA's were compared for their abilities to bind to elongation factor Tu (EF-Tu) in the presence both of GTP and guanylylimidodiphosphate (GMP-P(NH)P) . Formation of aminoacyl-tRNA . EF-Tu . guanine nucleotide ternary complexes was monitored by gel filtration on Sephadex G-100 and Ultrogel ACA 44 columns and also by measurement of the ability of the factor to diminish the rate of chemical hydrolysis of the aminoacyl-tRNA's . The apparent positional specificity of the factor was found to be affected substantially both by the choice of guanine nucleotide and gel filtration resin utilized, but not in any systematic fashion . Likewise, assay of ternary complex formation by diminution of the rate of chemical deacylation failed to reveal any consistent positional preference from one isoacceptor to another . It is worthy of note that each modified aminoacyl-tRNA tested did form a ternary complex with EF-Tu under each of the experimental conditions used for assay, but that in each case the difference in affinity of the factor for isomeric aminoacyl-tRNA's was less than that between either of the modified aminoacyl-tRNA's and the corresponding unmodified species . On the basis of the experiments performed, we conclude that (i) EF-Tu has remarkable conformation flexibility, possibly reflecting its physiological role in recognizing 20 tRNA isoacceptors and (ii) the factor has no obvious preference for a single positional isomer of aminoacyl-tRNA and it is not clear that any preference that might exist could be established convincingly using tRNA's terminating in 2'- and 3'-deoxyadenosine. J Biol Chem, 1979 Aug 10, 254(15), 6880 - 8 Hemoglobin switching in sheep . Synthesis, cloning, and characterization of DNA sequences coding for the beta B, beta C, and gamma-globin mRNAs; Benz EJ Jr et al.; Synthetic double-stranded DNAs (sDNAs) were prepared from sheep globin mRNA templates isolated from reticulocytes producing either hemoglobin B (HbB) (alpha 2 beta B2), HbC (alpha 2 beta C2), or HbF (alpha 2 gamma 2) . These DNAs were inserted into the Eco RI site of plasmid pMB9 by the homopolymer tailing method and used to transform Escherichia coli X1776 to tetracycline resistance . Recombinant clones were identified by colony hybridization and further characterized by molecular hybridization and restriction endonuclease analysis . All plasmids analyzed thus far contained either beta- or gamma-globin DNA sequences . Moreover, sDNAs used for cloning yielded restriction endonuclease fragments consistent with the presence of predominantly beta- or gamma-sDNA, indicating that formation of double-stranded alpha-sDNA proceeds much less efficiently under our conditions than the formation of non-alpha-sDNAs . Three recombinant plasmids, pS beta B2, pS beta C69, and pS gamma 56, were selected for detailed study . These were shown to contain, respectively, beta B-, beta C-, and gamma-DNA sequences by molecular hybridization and by protection of the appropriate cDNAs from S1 nuclease digestion . Each contained all of the restriction endonuclease sites defined for the synthetic sDNAs and protected at least 90% of the sequence length of homologous cDNA . Restriction endonuclease maps of the beta B- and beta C-globin genes were identical at all 12 sites that were mapped, whereas four differences were identified in the gamma gene compared to the two others; three of these corresponded to differences in amino acid sequence of the globins . A method was developed to isolate the anti-mRNA strand of the insert for use as a specific molecular hybridization probe analogous to complementary DNA. J Biol Chem, 1979 Aug 10, 254(15), 6873 - 5 Transfer RNA control of the activation of isomeric tRNATrp's; Alford BL et al.; Previous studies of the homologous aminoacylations of Escherichia coli and yeast tRNATrp's terminating in 2'- and 3'-deoxyadenosine established that E . coli tryptophanyl-tRNA synthetase activates its cognate tRNA preferentially on the 2' position, while the corresponding yeast enzyme utilizes the 3' position on its homologous substrate tRNA . As this seemed to be the only change in positional specificity during evolution, the heterologous activations were investigated in an effort to determine the basis for this change . Remarkably, E . coli tRNATrp terminating in 3'-deoxyadenosine was found to be the preferred substrate for both the E . coli and yeast activating enzymes, while the same tryptophanyl-tRNA synthetase preparations both activated the isomeric yeast tRNATrp's preferentially on the 3' position . Thus, the preferred position of activation was found to be specified by the tRNA rather than the activating enzyme and, additionally, to be due to some process not reflected in initial velocity measurements . The variable utilization of individual modified aminoacyl-tRNA's as substrates in an enzyme-catalyzed deacylation process appears to provide the most likely explanation for the experimental observations. J Biol Chem, 1979 Aug 10, 254(15), 7917 - 202 Control of membrane lipid synthesis in Escherichia coli during growth and during the stringent response; Snider MD; The regulation of phospholipid synthesis in cells of Escherichia coli was studied in vivo during growth and during the stringent response to amino acid starvation . Strains harboring the hybrid plasmid pLC44-14 (Clark, L., and Carbon, J . (1976) Cell 9, 91-99), which had increased levels of glycerophosphate acyltransferase, were used to study the involvement of this enzyme in the control of phospholipid synthesis . In addition, regulation was studied by measuring the levels of three early intermediates of phospholipid synthesis:phosphatidic acid, CDP-diglyceride, and dCDP-diglyceride . The liponucleotides were measured by a new enzymatic method which allows determinations to be made on crude lipid extracts . Results from experiments on growing cells are consistent with regulation of membrane lipid synthesis occurring in fatty acid synthesis or at the level of glycerophosphate acylation, but not at any later step . Experiments on the inhibition of lipid synthesis during the stringent response make it possible to rule out explanations which involve the inhibition of a single enzyme; enzymes both before and after the liponucleotides in phospholipid synthesis must be affected. Biochemistry, 1979 Aug 7, 18(16), 3557 - 63 Mechanism of action of D-serine dehydratase . Identification of a transient intermediate; Schnackerz KD et al.; Static absorbance measurements of D-serine dehydratase from Escherichia coli taken at 2 degrees C show that during the steady-state course of D-serine conversion the absorption maximum of the Schiff base of the cofactor pyridoxal 5'-phosphate (pyridoxal-P) is shifted from 415 to 442 nm . Furthermore, the progress curve of intermediates was monitored by stopped-flow techniques at wavelengths ranging from 320 to 500 nm . A point by point construction of successive spectra from these stopped-flow traces at various time intervals after the start of reaction resulted in a series of consecutive spectra exhibiting two isobestic points at 353 and 419 nm . The half-time of the absorbance changes occurring at 330 and 455 nm was found to be 6.5 ms, suggesting the observation of a single, enzyme-bound intermediate . The spectral data with substrate and inhibitors provide evidence that the intermediate is the Schiff base of alpha-aminoacrylate and pyridoxal-P . The proposed assignment is strongly supported by experiments of apodehydratase with transient-state analogues which exhibit a similar absorbance shift on binding to apoenzyme . Moreover, these results suggest that the phosphate group of the substrate--pyridoxal-P complex serves as the main anchoring point during catalysis . A reaction mechanism of the D-serine dehydratase is presented. Science, 1979 Aug 3, 205(4405), 508 - 11 A relationship between DNA helix stability and recognition sites for RNA polymerase; Vollenweider HJ et al.; The RNA polymerase binding sites on the DNA of (i) the aroE-trkA-spc segment of the Escherichia coli genome, (ii) transposon Tn3, (iii) plasmid ColE1, and (iv) coliphage lambda were mapped by electron microscopy, with the use of the BAC technique; these maps were compared with the maps of the early-melting regions for the same genomes . The results indicate that in all these cases the binding sites for the E . coli RNA polymerase lie preferentially in the early melting regions of DNA . These data indicate that helix stability may be an important feature of the multipartite nature of the promoter structure. Trop Anim Health Prod, 1979 Aug, 11(3), 171 - 4 Cell counts in bulked milk supplies from dairy farms of northern Nigeria; Lombin LH et al.; Somatic cells in 596 milk samples collected from bulked supplies of 3 northern Nigeria dairy farms were counted by an electronic method following a standard method for the preparation of the milk samples . Mean somatic cell counts per ml indicating low level of infection were 158,597, 166,742 and 155,032 for the 3 farms, without any significant differences . Mean somatic cell counts per ml indicating herd mastitis averaged 354,768 +/- 66,348 and the pathogenic organisms isolated were Escherichia coli and Staphylocucus aureus . Counts useful for future regular monitoring of somatic cells in bulked milk supplies in northern Nigeria are presented. J Cell Biol, 1979 Aug, 82(2), 369 - 79 Localization of submembranous cations to the leading end of human neutrophils during chemotaxis; Cramer EB et al.; Potassium pyroantimonate was used to localize sites of bound cations in human neutrophils under conditions of random migration, stimulated random migration (chemokinesis), and directed migration (chemotaxis) . The cells were placed in a standard chamber in which 0.45-micron micropore filters separated the cells from the stimulus (buffer, Escherichia coli endotoxin-activated serum or the synthetic chemotactic peptide N-formyl-Met-Leu-Phe) . The small pore filters permitted pseudopod formation but impeded cell imgration through the filter . Cells examined under all conditions had electron-dense precipitates of antimonate salts in some granules . However, antimonate deposits were localized in the condensed chromatin of the nucleus during random migration and associated to a large extent with the uncondensed nuclear chromatin during chemokinesis and chemotaxis . Under conditions of chemokinesis deposition of antimonate procipitates appeared on the cytoplasmic side of the plasma membrane of neutrophils whereas under conditions of chemotaxis cation deposits beneath the cell membrane were localized to the pseudopods which were directed toward the chemoattractant . In addition to endotoxin-activated serum, concentrations of N-formyl-Met-Leu-Phe which caused neutrophil chemotaxis (10(-8) M) also caused cation deposition beneath the cell membrane at the leading end of the cell regardless of whether albumin was present in the incubation media . However, with higher concentrations of the synthetic peptide (10(-5) M) which caused granule release and were not chemotactic, submembranous cation deposition was not seen . EDTA (10 mM) and EGTA (10 mM) removed nuclear, granular, and submembranous cation deposits from neutrophils examined under conditions of chemotaxis . X-ray microprobe analysis of antimonate deposits revealed the possible presence of calcium but did not detect sodium or magnesium . The data indicate that chemotactic factors induce submembranous deposition of cations, most likely Ca++, which localize to the leading edge of cells exposed to a gradient of chemoattractant. Med J Zambia, 1979 Aug-Sep, 13(4), 67 - 70 Tubo-ovarian abscess: a review of 46 cases; Chatterjee TK et al.; Forty-six patients with tubo-ovarian ascess are analysed . The abscess developed mostly in young multiparous women soon after menstruation . Coliforms were the main causative organism . The abscess resolved with conservative management in 21 cases and surgical intervention was necessary in 25 . The mortality (4.4%) due to conservative medical and surgical management was nearly half that of radical surgery. Can J Microbiol, 1979 Aug, 25(8), 937 - 9 A new gene involved in expression of fructose-1,6-diphosphate aldolase activity in Escherichia coli; Atherly AG et al.; A new gene, fdaB, has been mapped by transduction and partial diploid analyses and located adjacent to argA at 59.9 min on the Escherichia coli recalibrated linkage map . This gene is involved in expression of fructose-1,6-diphosphate aldolase activity and indirectly in ribosomal RNA synthesis . The temperature-sensitive mutant strain AA-157, containing the defective gene product of of fdaB, accumulates high concentrations of fructose 1,6-diphosphate at the nonpermissive temperature. Genetics, 1979 Aug, 92(4), 1041 - 59 Isolation and characterization of dnaX and dnaY temperature-sensitive mutants of Escherichia coli; Henson JM et al.; Escherichia coli mutants with temperature-sensitive (ts) mutations in dnaX and dnaY genes have been isolated . Based on transduction by phage P1, dnaX and Y have been mapped at minutes 10.4--10.5 and 12.1, respectively, in the sequence dnaX purE dnaY . Both dna Xts36 and Yts10 are recessive to wild-type alleles present on episomes . F13 carries both dnaX+ and Y+; the shorter F210 carries dnaY+, but not X+ . Lambda tranducing phages that carry dnaX+ or Y+ have been isolated, and hybrid plasmids of Col E1 and E . coli DNA from the Clarke and Carbon (1976) collection also carry portions of the dnaX purE dnaY region . Results obtained with the lambda transducing phages and the hybrid plasmids suggest that dnaX is a different gene from the previously characterized dnaZ gene, which is also near minute 10.5--The dnaXts36 mutant, after a shift to 42 degrees, stopped DNA synthesis gradually, and the total amount of DNA increased two-fold . When this mutant was shifted to 44 degrees, the rate of DNA synthesis dropped immediately and the final increment of DNA was only 10% of the initial amount . Replicative DNA synthesis in toluene-treated cells was completely inhibited at 42 degrees and was partially inhibited even at 30 degrees.--When the dnaYts10 mutant was shifted to 42 degrees, DNA synthesis gradually stopped, and the amount of DNA increased 3.6-fold . At 44 degrees, residual DNA synthesis amounted to a two-fold increase . Replicative DNA synthesis in vitro in toluene-treated cells was inactivated after 20 minutes at 42 degrees or by "preincubation" of cells at 42 degrees before toluene treatment.--The dnaX and dnaY products probably function in polymerization of DNA, although participation also in initiation cannot be excluded. Mol Gen Genet, 1979 Aug, 175(1), 53 - 6 IS2-43 and IS2-44: new alleles of the insertion sequence IS2 which have promoter activity; Sommer H et al.; The sequence of two new IS2 alleles with promoter activity (IS2-43 and IS2-44) is reported . The alleles are identical and are formed by a 17 bp tandem duplication in an AT-rich region of IS2 . This created a new RNA polymerase binding site . A mutation was found that increased the frequency of formation of these 17 bp duplications but not of another class of duplications, the "mini-insertions" . This suggested that the mechanisms of formation of the two classes of duplications are different. Mol Gen Genet, 1979 Aug, 175(1), 39 - 44 Construction and expression of a hybrid plasmid containing the Escherichia coli thrA and thrB genes; Cossart P et al.; In vitro recombination techniques were used to clone the Escherichia coli thrA and thrB structural genes in the plasmid vector pBR322 . The chimeric plasmid was analyzed and characterized genetically, by restriction mapping and DNA sequencing . The limited expression of the threonine biosynthetic enzymes in the strain carrying the recombinant plasmid is discussed. Mol Gen Genet, 1979 Aug, 175(1), 31 - 8 Regulation of dihydrofolate reductase synthesis in Escherichia coli; Smith DR et al.; Two clones from the Clarke-Carbon Escherichia coli colony bank were resistant to inhibition by trimethoprim, a potent inhibitor of dihydrofolate reductase . Both clones had elevated levels of dihydrofolate reductase . Furthermore, trimethoprim resistance and elevated enzyme levels were associated with ColE1 plasmids that carried DNA from the trkC ksgA pdxA region of the E . coli chromosome . Plasmid pLC1437a was shown by two criteria to carry the structural gene for dihydrofolate reductase: 1) A partial diploid containing plasmid pLC1437a produced a kinetically-recognizable dihydrofolate reductase that was not present in the parent haploid strain . 2) Plasmid pLC1437a coded for dihydrofolate reductase in vitro . A 1,000 base pair fragment of plasmid pLC1437a containing fol was used as a probe to measure fol mRNA in a mutant strain isolated by Sheldon and Brenner (Molec . gen . Genet . 147, 91-97, 1976) . The mutation in this strain, which results in constitutively-high levels of dihydrofolate reductase and in the inability of the strain to grow at 42 degrees C, is cis dominant (Sheldon and Brenner, 1976) . The results of kinetic hybridization and pulse-labeling experiments indicated that the regulatory mutant produced elevated levels of dihydrofolate reductase in response to an increased rate of synthesis of fol mRNA. Mol Gen Genet, 1979 Aug, 175(1), 13 - 7 Radiation sensitivity of messenger RNA; Ponta H et al.; Messenger RNA function is inactivated by irradiation with ultraviolet light . A unit length mRNA (in bases) is 2-3 times more sensitive than a unit length of DNA (in base pairs) with respect to the inactivation of template function . These data stem from four experimental systems all of which do not repair DNA: the translation of E . coli mRNA in rifampicin-treated cells, of T7 mRNA in infected E . coli, of f2 phage RNA in vivo, and of stable mRNA in chromosomeless minicells . The comparison of relative sensitivities to UV is relevant to the technique of UV mapping of transcription units which enjoys increasing popularity in pro- and eukaryotic genetic research. J Gen Microbiol, 1979 Aug, 113(2), 425 - 7 Specific proline accumulation in an acrA mutant of Escherichia coli K12 grown in salt-hypertonic medium; Nakamura H; Free proline is specifically accumulated in an acrA mutant of Escherichia coli K12 when cultured in a medium containing excess NaCl. J Gen Microbiol, 1979 Aug, 113(2), 297 - 303 Mapping of a new hem gene in Escherichia coli K12; Sasarman A et al.; A new type of haem-deficient mutant was isolated in Escherichia coli K12 by neomycin selection . The mutant, designated SASX38, accumulated uroporphyrin, coproporphyrin and protoporphyrin . Since it possessed normal ferrochelatase activity, it was assumed to be deficient in protoporphyrinogen oxidase activity . The gene affected in the mutant was designated hemG . Mapping of the hemG gene by phage P1-mediated transduction showed that it was located very close to the chlB gene (frequency of cotransduction 78.7%), between the metE and rha markers . This location is distinct from the other known hem loci in E . coli K12. Gann, 1979 Aug, 70(4), 421 - 8 Properties of interferon induced by purified protein derivative of tuberculin in mice sensitized with BCG or cell-wall skeleton of BCG; Takeyama H et al.; Mice sensitized with either BCG or cell-wall skeleton of BCG (BCG-CWS) produced interferon in blood after stimulation with specific antigen, purified protein derivative of tuberculin (PPD) . Both BCG-infected normal (C57BL/6) and thymic nude (BALB/c, nu/nu) mice showed enhanced activity to produce interferon by stimulation with E . coli endotoxin . However, detectable interferon was not produced in athymic nude mice sensitized with BCG or BCG-CWS by stimulation with PPD . Immune-induced interferon (I-IF) produced by BCG-CWS and PPD in mice was different in biological and physicochemical properties from virus-induced interferon . I-IF showed about 100 times more potent L-cell growth inhibitory activity than virus-induced L-cell interferon (L-IF) . Both I-IF and L-IF showed macrophage-activating activity, which renders resting macrphages cytotoxic to L1210 leukemia cells . Antiviral and macrophage-activating activity of interferon preparation was not separated physicochemically in this study. Biokhimiia, 1979 Aug, 44(8), 1377 - 80 {Kinetics of phenylacetamide hydrolysis by immobilized penicillinamidase}; Milman IA et al.; The kinetics of phenylacetamide hydrolysis catalyzed by polyacrylamide gel-immobilized penicillinamidase were studied . The Km and Kp values obtained were compared to the literary data for the specific substrate--benzylpenicillin . It was shown that the type of inhibition by the reaction product was the same, whereas the efficiency of binding of phenylacetic acid depended on the substrate structure. Agents Actions, 1979 Aug, 9(3), 280 - 3 The role of prostaglandins in experimental ocular inflammations; Yamauchi H et al.; Intraocular inflammations (uveitis) were produced in rabbits by intravitreal injection of killed and dried Mycobacterium butyricum or E . coli endotoxin, or paracentesis of the anterior chamber . The increase in permeability of the blood-aqueous barrier and the leucocyte migration into the aqueous humor were observed after these stimuli, although the leucocyte did not migrate after paracentesis . Topically applied indomethacin reduced these inflammatory parameters in the latter two models . However, ththacin, though the leucocyte was not affected by indomethacin, though the leucocyte migration was reduced . On the other hand, prostaglandin-like substances in thces of these substances were detected in the former model . These results indicated that, unlike the increase in the permeability of the blood-aqueous barrier after endotoxin injection andparacentesis, the response after M . butyricum injection is not mediated by prostaglandins . The role of prostaglandins in the leucocyte migration in these ocular inflammations was also discussed. Res Commun Chem Pathol Pharmacol, 1979 Aug, 25(2), 293 - 306 Studies on the mode of action of partially thiolated polycytidylic acid (MPC), a novel type of antineoplastic agent; Ho YK; Partially thiolated polycytidylic acid (MPC), a representative member of the "antitemplate" class of novel chemotherapeutic agents, is a potent inhibitor of the E . coli DNA-dependent RNA polymerase . It inhibited 50% of the enzymic reaction at a concentration of 6 micrometers . Kinetic studies indicated that MPC had no effect on the chain elongation of the transcription process, but it appeared to inhibit the initiation of RNA synthtesis presumably by competing with the DNA template for binding to the RNA polymerase . Binding studies, using a gel filtration method, showed that MPC and the RNA polymerase formed a stable complex which was not dissociated by 0.3 M NaCl . It is inferred that mixed disulfide linkage(s) might have been formed between the enzyme and MPC . The implications of these findings are discussed. Proc Natl Acad Sci U S A, 1979 Aug, 76(8), 4020 - 4 Identification and characterization of a self-regulated repressor of translocation of the Tn3 element; Chou J et al.; Gene fusions that bring expression of the lacZ gene under control of transcriptional and trnaslational signals within the transposable element Tn3 have been used to study regulation of Tn3-specified proteins . A gene encoding a 21,355-Mr peptide that represses translocation of Tn3 and acts at the level oquenced; amber, missense, and cis-dominant (operator-constitutive) point mutations in this gene have been isolated and characterized. Proc Natl Acad Sci U S A, 1979 Aug, 76(8), 3795 - 9 Reconstitution of RNase P activity from inactive RNA and protein; Kole R et al.; RNase P preparations from Escherichia coli can be separated into RNA and protein by chromatography, in buffers containing 7 M urea, on Sephadex G-200, DEAE-Sephadex, or CM-Sephadex columns . Neither RNA nor protein components alone exhibits any RNase activity . RNase P activity can be reconstituted by mixing separated RNA and protein components in buffer containing 7M urea followed by dialysis of this mixture to remove the urea . Of several purified RNAs tried, only M2 RNA, the RNA species found in purified RNase P, is active in the reconstitution experiments. Proc Natl Acad Sci U S A, 1979 Aug, 76(8), 3769 - 73 Ribosome structure: localization of 3' end of RNA in small subunit by immunoelectronmicroscopy; Olson HM et al.; The 3' end of the RNA in the 30S ribosomal subunit of Escherichia coli has been modified by oxidation with sodium periodate and conjugation with the (mono) dinitrophenyl derivative of ethylenediamine . Antibodies, induced with dinitrophenyl-bovine serum albumin, interact with the modified ribosomal subunits . Electron micrographs of negatively stained antibody-subunit complexes show individual ribosomal subunits to which a single antibody molecule is bound and subunit dimers cross-linked by an IgG molecule . The modified 3' terminus has been localized to a single site on the upper portion of the platform region of the 30S subunit . This location is consistent with earlier placements of proteins that react with the 3' end of the RNA. Proc Natl Acad Sci U S A, 1979 Aug, 76(8), 3632 - 6 A DNA primase specified by I-like plasmids; Lanka E et al.; An enzyme has been isolated from Escherichia coli strains harboring the I-like plasmid R64drd11, which is capable of initiating DNA synthesis on the circular, single-stranded DNA of phages phi X174, fd, and G4 . In the conversion of these templates to duplex forms in vitro, the enzyme can substitute for the functions of E . coli dna B-dnaB-dnaC-dnaG proteins, E . coli RNA polymerase, and E . coli dnaG protein, respectively . The enzyme requires all four ribonucleoside triphosphates for optimal activity, although a combination of ATP, CTP, and GTP can almost completely satisfy the rNTP requirement . The enzyme appears to cooperate specifically with DNA polymerases III because single-stranded DNA-dependent synthesis takes place in extracts deficient in DNA polymerases I and II but not in extracts from a dnaZ mutant . Highly purified enzyme preparations consist mostly of two major polypeptides, Mr 140,000 and 180,000, when analyzed by sodium dodecyl sulfate gel electrophoresis . These polypeptides cosediment with the enzyme activity through a glycerol gradient with a sedimentation coefficient of 3.6 S . DNA priming activity in extracts of E . coli strains harboring the mutant plasmids R64drd11 or ColIdrd1, which are derepressed in functions of conjugational DNA transfer, severalfold higher than the activity from strains carrying the corresponding wild-type plasmid . This correlation suggests that the enzyme may play a role in conjugational DNA synthesis. Mutat Res, 1979 Aug, 62(1), 7 - 17 A recA-dependent mutator of Escherichia coli K12: method of isolation and initial characterization; Hombrecher G et al.; A number of mutator strains of E . coli were isolated using histochemical techniques which allow the identification of a single mutator colony on agar plates with as many as 2000 colonies . Several mutators isolated in this way were found by P1-mediated transduction to map to the proA--proB region of the E . coli chromosome . The map position of these mutators is very close to that of the conditional mutator, mutD . However, in contrast to mutD, one of these newly isolated mutators was suppressed in a thermosensitive recA strain at 43 degrees C, but not at 30 degrees C . This mutator mutation has been named mut-8 . Besides being dependent upon recA, mut-8 is also dependent upon growth in enriched medium for the expression of its mutator activity . The mutator activity of mut-8 was found to be recessive to the wild-type allele. Infect Immun, 1979 Aug, 25(2), 603 - 9 Role of Escherichia coli K capsular antigens during complement activation, C3 fixation, and opsonization; Van Dijk WC et al.; Escherichia coli strains with K capsular polysaccharides are relatively resistant to phagocytosis by polymorphonuclear leukocytes, in contrast to E . coli strains without K antigens . This inhibition of phagocytosis is related to an impaired recognition of the K+ strains by the phagocytes due to ineffective opsonization . All five strains without K antigens were readily phagocytized after opsonization in 5% normal serum, compared with no uptake of the K+ strains . Evidence is presented that the decreased opsonization of the K+ strains in normal serum is caused by a low rate of complement activation of the strains, with subsequent absence of C3b fixation or C3d fixation or both to the cell wall of the bacteria . After removal of the K+ antigens by heating of a K+ E . coli strain, the strain was able to activate complement, to bind C3b or C3d or both, and to become opsonized . Complement was then activated via the classical and alternative pathways, which was comparable to the complement consumption by K- E . coli. Eur J Biochem, 1979 Aug 1, 98(2), 567 - 71 Reconstitution of active 30-S ribosomal subunits in vitro using heat-denatured 16-S rRNA; Barritault D et al.; 30-S ribosomal subunits which have been reconstituted using heat-denatured 16-S rRNA can participate in the synthesis of lysosyme in vitro . Therefore all the information contributed by 16-S rRNA to the reconstitution process is carried in the primary sequence of this RNA . The specific protein-synthesizing activity of 30-S subunits reconstituted from 30-S subunit proteins and heat-denatured 16-S rRNA is about one third of that observed if unheated 16-S rRNA is used and is comparable to the activity of 30-S particles isolated after dissociation of 70-S ribosomes in the presence of 0.1 mM Mg2+. Eur J Biochem, 1979 Aug 1, 98(2), 557 - 66 Protein . nucleic-acid reaction kinetics . Theoretical analysis of the binding reaction between DNA and RNA polymerase; Giacomoni PU; This paper presents methods developed in order to analyze experimental results concerning the binding of Escherichia coli DNA-dependent RNA polymerase to DNA at high and at low DNA concentrations, using the filter retention assay . The basis hypotheses, under which the mathematical expressions for describing the kinetics of binding are derived, are as follows . (a) At low DNA concentration: equivalence and independence of the specific binding sites; first-order dependence of the binding reaction on both DNA and protein concentration . (b) At high DNA concentration: equivalence and independence of the non-specific binding sites; no direct transfer or one-dimensional sliding of the protein along the DNA . Comparison between theoretical predictions and experimental results at high DNA concentration will allow one to determine the relative value of the rates of binding of RNA polymerase to different promoters (between 1 and 2 in T5 DNA) . Binding experiments performed at low DNA concentration are reported in this paper: these results and the analysis which is reported allow one to determine the value of the rate constant of formation of non-filterable complexes for the system fd DNA (replicative form) . RNA-polymerase (kappa a = 3.3 X 10(8) M-1 s-1 in 0.1 M NaCl, 0.01 M MgCl2). Mutat Res, 1979 Aug, 64(4), 241 - 8 Methodology for the testing of food dyes for genotoxic activity: experiments with red 2G (C.I . 18050); Haveland-Smith RB et al.; A methodology for investigating genotoxicity of food colours using the fluctuation and DNA-repair assays with bacteria is