J Bacteriol, 1981 Nov, 148(2), 720 - 3 Temperature-sensitive mutation affecting synthesis of phosphoenolpyruvate carboxykinase in Escherichia coli; Goldie AH et al.; A mutation has been characterized in Escherichia coli which results in temperature-sensitive expression of phosphoenolpyruvate carboxykinase activity and antigen . The enzyme produced by the mutant strain at a permissive temperature or by cells treated with chloramphenicol at nonpermissive temperatures had normal activity and stability in extracts . Since phosphoenolpyruvate carboxykinase had a monomeric structure, the mutation probably affects the synthesis, rather than the structure or assembly, of the enzyme. J Bacteriol, 1981 Nov, 148(2), 684 - 96 Comparison among patterns of macromolecular synthesis in Escherichia coli B/r at growth rates of less and more than one doubling per hour at 37 degrees C; Pierucci O et al.; In Escherichia coli B/r, the relationship between the patterns of chromosome replication and of synthesis of envelope components differs at various growth rates . At growth rates greater than 1.0 doubling per h at 37 degrees C, the average mass and age at initiation of rounds of chromosome replication are similar to those at increase in incorporation of precursors into a major outer membrane protein and phosphatidylethanolamine . At growth rates less than 1.0 doubling per h at 37 degrees C the average mass and age at increase in the synthesis of these envelope components differ from those at initiation of chromosome replication . The average cell mass per chromosomal origin at initiation of rounds of chromosome replication is not a constant and varies between growth rates greater and less than 1.0 doubling per h. J Bacteriol, 1981 Nov, 148(2), 551 - 8 New mechanism for post-translational processing during assembly of a cytoplasmic membrane protein? MacGregor CH, McElhaney GE. Insertion of nitrate reductase into the Escherichia coli cytoplasmic membrane was examined by following the fate of pulse-labeled enzyme in both the membrane and cytoplasm during various times after the addition of an unlabeled chase . The polypeptide composition of this labeled enzyme was determined by autoradiography of immunoprecipitated material after separation on sodium dodecyl sulfate-polyacrylamide gels . The data presented here indicate that immediately after appropriate insertion of the enzyme into the membrane, a post-translational event occurs which converts the cytoplasmically synthesized form of subunit B (B') to the form found in the completely assembled enzyme (B) . B' is distinguished from B by its more rapid electrophoretic mobility . B' was found in the cytoplasm of all strains tested, in the membrane of strains with defects in enzyme insertion (hemA and chlE), and as a transient component in the membrane of wild-type cells. J Bacteriol, 1981 Nov, 148(2), 472 - 9 L-arabinose transport systems in Escherichia coli K-12; Kolodrubetz D et al.; Mutations in the arabinose transport operons of Escherichia coli K-12 were isolated with the Mu lac phage by screening for cells in which beta-galactosidase is induced in the presence of L-arabinose . Standard genetic techniques were then used to isolate numerous mutations in either of the two transport systems . Complementation tests revealed only one gene, araE, in the low-affinity arabinose uptake system . P1 transduction placed araE between lysA (60.9 min) and thyA (60.5 min) and closer to lysA . The operon of the high-affinity transport system was found to contain two genes: araF, which codes for the arabinose-binding protein, and a new gene, araG . The newly identified gene, araG, was shown by two-dimensional gel electrophoresis to encode a protein which is located in the membrane . Only defects in araG could abolish uptake by the high-affinity system under the conditions we used. Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 7059 - 63 Transmission electron microscopic method for gene mapping on polytene chromosomes by in situ hybridization; Wu M et al.; A transmission electron microscope method for gene mapping by in situ hybridization to Drosophila polytene chromosomes has been developed . As electron-opaque labels, we use colloidal gold spheres having a diameter of 25 nm . The spheres are coated with a layer of protein to which Escherichia coli single-stranded DNA is photochemically crosslinked . Poly(dT) tails are added to the 3' OH ends of these DNA strands, and poly(dA) tails are added to the 3' OH ends of a fragmented cloned Drosophila DNA . These probe--dA strands are hybridized in situ to polytene chromosome squashes . Gold spheres are linked to the hybridized probe--dA strands by A.T base pairing . The sphere positions relative to the chromosome bands can be observed by transmission electron microscopy . The method shows low background and high resolution. Biochem J, 1981 Nov 1, 199(2), 383 - 92 Kinetic studies of the fatty acid synthetase multienzyme complex from Euglena gracilis variety bacillaris; Walker TA et al.; A fatty acid synthetase multienzyme complex was purified from Euglena gracilis variety bacillaris . The fatty acid synthetase activity is specifically inhibited by antibodies against Escherichia coli acyl-carrier protein . The Euglena enzyme system requires both NADPH and NADH for maximal activity . An analysis was done of the steady-state kinetics of the reaction catalysed by the fatty acid synthetase multienzyme complex . Initial-velocity studies were done in which the concentrations of the following pairs of substrates were varied: malonyl-CoA and acetyl-CoA, NADPH and acetyl-CoA, malonyl-CoA and NADPH . In all three cases patterns of the Ping Pong type were obtained . Product-inhibition studies were done with NADP+ and CoA . NADP+ is a competitive inhibitor with respect to NADPH, and uncompetitive with respect to malonyl-CoA and acetyl-CoA . CoA is uncompetitive with respect to NADPH and competitive with respect to malonyl-CoA and acetyl-CoA . When the concentrations of acetyl-CoA and malonyl-CoA were varied over a wide range, mutual competitive substrate inhibition was observed . When the fatty acid synthetase was incubated with radiolabelled acetyl-CoA or malonyl-CoA, labelled acyl-enzyme was isolated . The results are consistent with the idea that fatty acid synthesis proceeds by a multisite substituted-enzyme mechanism involving Ping Pong reactions at the following enzyme sites: acetyl transacylase, malonyl transacylase, beta-oxo acyl-enzyme synthetase and fatty acyl transacylase. J Virol, 1981 Nov, 40(2), 396 - 402 phi X174-directed DNA and protein syntheses in infected minicells; Reeve JN; Phi X174-infected minicells, produced by Escherichia coli PC2251, synthesized 11 phi X174-encoded polypeptides . The infecting single-stranded viral genome was converted to a double-stranded, closed circular, replicative form (replicative form I) . Little, if any, replicative form I replication took place, and synthesis of progeny single-stranded molecules could not be detected. J Virol, 1981 Nov, 40(2), 341 - 9 Coliphage 186 Replication is delayed when the host cell is UV irradiated before infection; Hooper I et al.; In contrast to results with injections by lambda and P2, the latent period for infection by coliphage 186 is extended when the host cell is UV irradiated before infection . We find that 186 replication is significantly delayed in such a cell, even though the phage itself has not been irradiated . In contrast, replication of the closely related phage P2 under the same conditions is not affected. J Gen Microbiol, 1981 Nov, 127, pt 1, 93 - 102 TnA-directed deletion of the trp operon from RSF2124-trp in Escherichia coli; Tsunekawa H et al.; Plasmid pMT-trp was constructed by digestion of RSF2124-trp with restriction endonuclease PstI and ligation with T4 ligase . In pMT-trp about 78% of the DNA of transposon TnA from RSF2124-trp was deleted, and hence the gene for ampicillin resistance was lost . All Trp- segregants from pMT-trp carriers in Escherichia coli W3110 and its derivatives were found to have lost the entire plasmid . On the other hand, deletion plasmids which had lost the trp operon were found among Trp- segregants from RSF2124-trp carriers, particularly from the mutant strain trpAE1 trpR tnaA . The experimental fact that deletion occurred exclusively in RSF2124-trp suggests that the presence of TnA in the plasmid (RSF2124-trp) was responsible for the deletion. Ann Microbiol (Paris), 1981 Nov-Dec, 132 B(3), 387 - 98 Suspected epidemiological relationships among strains of Escherichia coli O55:B5 confirmed by restriction pattern analysis of the carried plasmid (RColBM IncFIII); Sasarman A et al.; Seven strains of Escherichia coli O55:B5g, isolated from independent cases of infantile diarrhoea, contained similar RColBM plasmids beloning to incompatibility group IncFIII . Suspecting an epidemiological link among all these cases we compared the restriction pattern of the corresponding RColBM plasmids after digestion with endonucleases EcoRI and BglII . These patterns were similar, confirming the suspected relationships among the seven cases of infantile diarrhoea . The patterns obtained with the RColBM plasmids from children were different from those obtained with two RColBM plasmids isolated from adults . The probable source of the seven cases of infantile diarrhoea was identified as a nursery. Mol Biol (Mosk), 1981 Nov-Dec, 15(6), 1245 - 57 {Interaction of RNA polymerase with hybrid plasmids carrying Escherichia coli threonine operon}; Kozlov IuI et al.; The RNA polymerase binding with two hybrid plasmids carrying threonine operon genes was studied . RNA polymerase binding sites were localized using nitrocellulose binding assay and electron microscopy visualization of the RNA polymerase--DNA complexes . To confirm that observed RNA polymerase binding sites are real promoters we analyzed RNA transcripts synthesized in vitro by hybridization with DNA fragments . The promoters of following genes were localized on the plasmid maps: ampicilline, threonine and RNA-primer of DNA replication . Two latter promoters bound RNA polymerase only being in the supercoiled DNA molecule . Several additional binding sites were found . The positions of some of these sites corresponded with known sites of rho-independent transcription termination. Mol Biol (Mosk), 1981 Nov-Dec, 15(6), 1205 - 23 {Pyrophosphate analogs in the pyrophosphorolysis reaction catalyzed by Escherichia coli RNA polymerase}; Rozovskaia TA et al.; Processive pyrophosphorolysis of RNA from ternary RNA polymerase-nascent RNA-delta D111 T7 DNA complex has been followed in the absence of nucleoside triphosphates . Series of inorganic pyrophosphate analogs were investigated for their ability to sustain the reaction and to compete with inorganic pyrophosphate for the reaction . Methylenediphosphonic, imidodiphosphonic, phosphonacetic acids, inorganic triphosphate, methylenediphosphonic and phosphate were found to be capable of substituting the inorganic pyrophosphate in RNA degradation reaction with tantamount efficiency . They give rise to nucleoside monophosphates for phosphonoacetic acid, nucleoside triphosphates for inorganic pyrophosphate and inorganic triphosphate, nucleoside triphosphates analogs for methylenediphosphonic, imidodiphosphonic acids and methylenediphosphonic acid phosphate as the low molecular weight product of the reaction . The problem of specific interaction of RNA polymerase with nucleoside triphosphates and inorganic pyrophosphate is discussed in the terms of structural requirements for the compounds to be a potent substrate for RNA polymerase. J Virol, 1981 Nov, 40(2), 450 - 5 Vertebrate DNAs contain nucleotide sequences related to the transforming gene of avian myeloblastosis virus; Bergmann DG et al.; Avian myeloblastosis virus contains a continuous sequence of approximately 1,000 nucleotides which may represent a gene (amv) responsible for acute myeloblastic leukemia in chickens . This sequence appears to have been acquired from chicken DNA and to be substituted for the envelope gene in the viral genome . We used hybridization probes enriched for the amv sequences and conditions that facilitate annealing of partially homologous nucleotide sequences to show that cellular sequences related to amv are present in the genomes of all vertebrates ranging from amphibians to humans but were not detected in fish, sea urchins, or Escherichia coli . In contrast to the preceding findings, nontransforming endogenous proviral nucleotide sequences closely related to the remainder of the avian myeloblastosis virus genome and to the entire myeloblastosis-associated helper virus are present only in chicken DNA . The amv-related cellular sequences appear to be highly conserved during evolution and to be contained at only one or a few locations in the genome of vertebrates . Within closely related species, they appear to share common evolutionary genetic loci . These findings and similar ones obtained with other highly oncogenic retroviruses containing a transforming gene suggest a general mechanism for acquisition of viral oncogenic sequences and an essential role for these sequences in the normal cellular state. Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 7069 - 72 Inversions between ribosomal RNA genes of Escherichia coli; Hill CW et al.; It might be anticipated that the presence of redundant but oppositely oriented sequences in a chromosome could allow inversion of the intervening material through homologous recombination . For example, the ribosomal RNA gene rrnD of Escherichia coli has the opposite orientation fro rrnB and rrnE and is separated from these genes by roughly 20% of the chromosome . Starting with a derivative of Cavalli Hfr, we have constructed mutants that have an inversion of the segment between rrnD and either rrnB or rrnE . These mutants are generally quite viable but do exhibit a slight reduction in growth rate relative to the parental strain . A major line of laboratory E . coli, W3110 and its derivatives, also has an inversion between rrnD and rrnE, probably created directly by a recombinational event between these highly homologous genes. Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 6817 - 20 Base insertion and deletion mutations induced in an Escherichia coli plasmid by benzo{a}pyrene-7,8-dihydrodiol-9,10-epoxide; Mizusawa H et al.; Mutations induced by (+/-)trans-benzo-{a}pyrene-7,8-dihydrodiol-9,10-epoxide (BaP-diol epoxide-1) were selected by using a recombinant plasmid vector system designed for the study of transcription termination signals . The plasmid contains a transcription terminator positioned between a promotor signal and the Escherichia coli galactokinase structural gene (galK) . By selections for the expression of galK (i.e., galK- to galK+), mutations are obtained in the terminator region that allow transcription from the promotor to read the galK gene . These mutations were characterized by direct DNA sequence of the terminator region . The DNA sequence changes caused by BaP-diol epoxide-1 were demonstrated for three different mutants . Two were found to be single-base-pair insertions of T.A into a cluster of consecutive T.A base pairs and the other change was a single-base-pair deletion of G.C from a cluster of consecutive G.C base pairs. Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 6684 - 8 Molecular cloning of equine herpesvirus type 1 DNA: analysis of standard and defective viral genomes and viral sequences in oncogenically transformed cells; Robinson RA et al.; Genomic DNA sequences of equine herpesvirus type 1 (EHV-1) have been cloned as BamHI and EcoRI restriction fragments into the plasmid pBR322 and propagated in Escherichia coli . With the exception of two EcoRI restriction fragments that reside in the S region of the viral genome, all of the cloned fragments demonstrated the same electrophoretic mobilities, restriction cleavage sites, and blot-hybridization patterns as did the parent fragments produced by BamHI or EcoRI digestion of virion DNA . The EcoRI J fragment and the BamHI E fragment of the L-region terminus were cloned after the addition of appropriate linker oligonucleotides . Fragments originating from each of the two isomeric forms of EHV-1 DNA were contained in this library of clones . Supramolar DNA fragments present only in the DNA of defective interfering (DI) particles of EHV-1 were generated from Bgl II digestion of DNA preparations enriched for EHV-1 DI particles and were cloned as Bgl II and EcoRI fragments into the plasmid vector . The cloned viral sequences represented in this defective genome mapped to the S region of EHV-1 DNA . Blot-hybridization analysis of EHV-1 transformed and tumor cell DNAs with the cloned BamHI B fragment confirmed that subgenomic viral sequences are present and indicated that those sequences map to the viral genome between 0.32 and 0.43 map unit. Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 6670 - 4 Conditional expression of the vesicular stomatitis virus glycoprotein gene in Escherichia coli; Rose JK et al.; Bacterial plasmids that directed expression of the vesicular stomatitis virus glycoprotein (G-protein) gene under control of the tryptophan operon regulatory region were constructed . A plasmid directing the synthesis of a G-protein-like protein (containing the NH2-terminal segment of seven amino acids encoded by the trpE gene fused to the complete G-protein sequence lacking only its NH2-terminal methionine) could be transformed into trpR+ (repressed) but not into trpR- (derepressed) cells . This result suggested initially that derepressed synthesis of the G-protein-like protein encoded by this plasmid was lethal in Escherichia coli . Deletion of the sequence encoding the large hydrophobic segment near the COOH terminus of G-protein did not overcome this lethality . Lethality of derepressed synthesis was overcome by deletion of the G-protein gene region encoding 10 amino acids in the hydrophobic NH2-terminal domain (signal peptide) . Tryptic peptide mapping demonstrated that the G-protein-like protein and some truncated proteins encoded by the plasmid contained G-protein protein sequences . Antisera to vesicular stomatitis virus precipitated the G-protein-like protein, showing that it shares antigenic determinants with the authentic G-protein protein. J Bacteriol, 1981 Nov, 148(2), 521 - 6 Regulation of fatty acid degradation in Escherichia coli: analysis by operon fusion; Clark D; Fusion of the lacZ gene coding for beta-galactosidase to the fadA,B and fadE operons was accomplished by using the phage Mu d (Apr lac) . In such fusion strains, beta-galactosidase was induced by long-chain fatty acids and repressed by glucose, as is the normal pattern of control for the enzymes of the fad regulon . The level of induction seen was approximately 10-fold for both the fadA and fadE operons . These results demonstrate that the previously observed regulation of both the fadA and fadE operons is at the transcriptional level . When an insertion mutation in the fadR (repressor) gene was introduced into the fusion strains, beta-galactosidase was produced constitutively . A series of fatty acids of different chain lengths were tested as inducers . Acids of chain lengths of 10 carbon atoms or less failed to induce, those of 12 carbon atoms induced partly, and those of 14 or more carbon atoms induced fully . Imidazole was found to counteract the glucose repression of the fadA operon as recently demonstrated for the ara operon. J Bacteriol, 1981 Nov, 148(2), 498 - 507 Cloning of immunity and structural genes for colicin V; Frick KK et al.; The colicin V immunity and structural genes of plasmid pColV-B188 were cloned into the vectors pMB9, pBR322, and pMK16 . Both genes are closely linked and can be isolated on a 900-base-pair deoxyribonucleic acid fragment . Insertion of the transposon Tn5 into this cloned sequence led to the construction of a mutant plasmid which conferred colicin V immunity, but not the ability to produce this colicin . Analysis of the products determined by these cloned genes in cells has led to the conclusion that the polypeptide involved in immunity has a molecular weight of about 6,500, whereas the colicin has a molecular weight of approximately 4,000. J Bacteriol, 1981 Nov, 148(2), 450 - 8 New nalidixic acid resistance mutations related to deoxyribonucleic acid gyrase activity; Yamagishi J et al.; In Escherichia coli K-12 mutants which had a new nalidixic acid resistance mutation at about 82 min on the chromosome map, cell growth was resistant to or hypersusceptible to nalidixic acid, oxolinic acid, piromidic acid, pipemidic acid, and novobiocin . Deoxyribonucleic acid gyrase activity as tested by supercoiling of lambda phage deoxyribonucleic acid inside the mutants was similarly resistant or hypersusceptible to the compounds . The drug concentrations required for gyrase inhibition were much higher than those for cell growth inhibition but similar to those for inhibition of lambda phage multiplication . Transduction analysis with lambda phages carrying the chromosomal fragment of the tnaA-gyrB region suggested that one of the mutations, nal-31, was located on the gyrB gene. Invest Urol, 1981 Nov, 19(3), 148 - 53 Immunology of pyelonephritis in the primate model . II . Effect on immunosuppression; Roberts JA et al.; Nonobstructive pyelonephritis was produced in the rhesus monkey (Macaca mulatta) by means of retrograde inoculation of Escherichia coli to the point of pyelotubular backflow . The effects of treatment with cyclophosphamide or azathioprine were determined . Commensal renal viruses were not activated by the immunosuppression, and thus did not complicate our results . Both cyclophosphamide and azathioprine prolonged the bacteriuria and produced more severe pathology . Cyclophosphamide decreased the leukocytic response and partically suppressed the antibody response . The increased amount of acute pyelonecytic response and partially suppressed the antibody response . The increased amount of acute pyelonephritic lesions after treatment with this drug suggest that the antibody and inflammatory responses may be important protective mechanisms, particularly regarding prevention of abscesses . In contrast, azathioprine did not decrease the leukocytosis nor the antibody responses, but resulted in decreased in vitro responsiveness of lymphocytes to mitogens . The increased severity of chronic pyelonephritic lesions after azathioprine treatment suggests that the cellular immune response also may be an important protective mechanism during late stages of the disease . The results thus indicate that the immune response is protective and is not directly responsible for the chronic scarring of pyelonephritis. Gene, 1981 Nov, 15(2-3), 151 - 6 DNA sequence of the att region of coliphage 434; Mascarenhas D et al.; Phages lambda and 434 are related phages that insert at the same site on the Escherichia coli chromosome . A 5.9-kb SalI-BamHI fragment derived from phage 434 was shown to hybridize to a 0.5-kb probe carrying attP-lambda . A 0.8-kb Bam HI-TaqI fragment subcloned into pBR327 was used for sequencing . The sequence of the 500 bp around the insertion site is given here, Comparison of the lambda and 434 sequence shows that the following regions are conserved: the coding sequence for the integrase protein (only 162 bp have been sequenced corresponding to the carboxy terminus), the 15-bp common core at the insertion site, and the three integrase-binding sites flanking the insertion site . The lambda and 434 sequences diverge radically to the left of base-197, suggesting that DNA to the left of that point plays no specific role in insertion or its regulation. Gene, 1981 Nov, 15(2-3), 139 - 49 Transposition of a tetracycline-resistance determinant (Tn1523) and cointegration events mediated by the pIP231 plasmid in Escherichia coli; Briaux-Gerbaud S et al.; A 10.8-kb transposable DNA sequence conferring resistance to tetracycline resides on the IncY Escherichia coli plasmid pIP231 . This sequence, designated Tn1523, was shown to insert into different sites of the replicons of the IncY prophage P1Cm c1.100 and the IncI1 plasmid pIP112 . This process is not dependent on the host recombination system recA . Genetic results indicate that Tn1523 transposition involves the formation of a cointegrate intermediate, either between pIP231 and P1Cm c1.100, or between pIP231 and pIP112 . These intermediates were found to be resolved into donor and recipient plasmids, each harboring a copy of the Tn1523 transposon . A stable structure formed by fusion of the pIP231 plasmid with the pIP112 plasmid was also observed . This event occurs in the absence of the bacterial recA gene product and seems to involve a site-specific reciprocal recombination between "IS-like" elements. Gene, 1981 Nov, 15(2-3), 119 - 26 The nucleotide sequence surrounding the promoter region of colicin E1 gene; Ebina Y et al.; The nucleotide sequence of 570 bp, covering the N-terminal portion of the colicin E1 gene, was determined . The sequence of the N-terminal four amino acids of the colicin E1 protein, determined by manual Edman degradation, agreed with that predicted from the nucleotide sequence . From analysis of the 5'-terminal sequences of RNAs synthesized in vitro, the promoter and operator regions of the colicin E1 gene were assigned . These data indicate the existence of two promoters, one of which is located in the coding region for colicin E1 . DNA sequence homology of 16 bp was found between the putative operator regions of the colicin E1 and recA genes. Rev Infect Dis, 1981 Nov-Dec, 3(6), 1186 - 95 Isolation and characterization of a human fibroblast interferon gene and its expression in Escherichia coli; Derynck R et al.; Human fibroblasts were induced for interferon synthesis, and the mRNA coding for interferon was partially purified . On this basis, double-stranded DNA copies were synthesized enzymatically and were inserted into cloning vehicles . A large collection of colonies containing chimeric plasmids was obtained . An RNA selection method was used for the identification of several individual clones containing the human fibroblast interferon (IFN-beta) cDNA . From the nucleotide sequence of the cloned structural gene, the primary structure of the mRNA and, hence, of the protein itself was deduced . IFN-beta is a polypeptide 166 amino acids long and contains a single site for N-glycosylation; IFM-beta is normally made as a preinterferon containing a signal sequence that is 21 amino acids long . The interferon gene was inserted into an expression plasmid under thermoinducible control of the phage lambda PL promoter . this allowed the synthesis of polypeptides in the bacteria, which for all physicochemical, biological, and immunologic properties tested closely resembled authentic IFN-beta. Mol Biol (Mosk), 1981 Nov-Dec, 15(6), 1224 - 33 {Enzymatic synthesis and molecular cloning of the pigeon alpha-globin structural gene}; Skobeleva NA et al.; Double-stranded DNA synthesized from the pigeon globin mRNA by the subsequent actions of avian myeloblastosis virus reverse transcriptase and E . coli DNA polymerase I was split with nuclease S1 and inserted into PstI site in the plasmid pBR322 by poly(dG) times poly(dC) homopolymer extension technique using terminal deoxynucleotidyl transferase . E . coli transformants have been shown to contain pigeon globin sequences by colony hybridization with pigeon globin {32P}cDNA . The inserted DNA fragment deleted from recombinant DNA by PstI treatment hybridizes with globin cDNA . The maximal length of the inserted fragment measured in agarose gel was found to be 550--560 base pairs . Inserted sequences subjected to analysis by hybridization with alpha- and beta-{32P}cDNA have been ascribed to the pigeon alpha globin chain . EcoRI, HindIII, BglII, SalI, BamHI, PstI restriction enzymes did not cleave the inserted DNA fragment . Pigeon DNA coding alpha-globin chain contains recognition sites for AluI, HindII and HaeIII restriction enzymes . Part of the recombinant clones remains resistant to ampicillin and therefore in some of these clones the globin gene could be expressed. Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 6613 - 7 Use of synthetic oligonucleotides as hybridization probes: isolation of cloned cDNA sequences for human beta 2-microglobulin; Suggs SV et al.; We have synthesized two sets of 15-base-long oligodeoxyribonucleotides corresponding to all possible coding sequences for a small portion of human beta 2-microglobulin . Labeled oligonucleotides were used as hybridization probes to screen bacterial clones containing cDNA sequences primed with oligo(dT) and inserted into the plasmid vector pBR322 . One beta 2-microglobulin cDNA clone was detected in the 535 bacterial plasmid clones that were screened . The clone has been characterized by blotting and nucleotide sequence analysis . The cloned beta 2-microglobulin sequence contains 217 base pairs of the 3' untranslated region of the mRNA and 328 base pairs (97%) of the coding region. Eur J Biochem, 1981 Nov, 120(1), 197 - 202 Synthesis of human interferon beta 1 in Escherichia coli infected by a lambda phage recombinant containing a human genomic fragment; Mory Y et al.; DNA from a human adult was fragmented by partial digestion with restriction endonuclease EcoRI and cloned in lambda Charon 4A . Clone C15, with a human DNA insert of 17 X 10(3) bases, was identified as containing a gene for the fibroblast interferon, interferon beta 1 . Restriction mapping shows that this gene, located on a 1840-base EcoRI fragment, is not interrupted by introns . Moreover, we show that this human genomic DNA fragment is able to direct the synthesis of active human interferon beta 1 in Escherichia coli . Interferon activity of up to 7 X 10(6) U/l was recovered from phage lysates by chromatography on Cibacron blue--Sepharose, and had the same immunological properties and species specificity as interferon produced by human fibroblasts. Eur J Biochem, 1981 Nov, 120(1), 1 - 7 Cloning of four DNA fragments complementary to human thyroglobulin messenger RNA; Berge-Lefranc JL et al.; Human thyroglobulin mRNA was isolated from Graves' goitres by size selection of total poly(A)-rich RNA in a sucrose gradient . It sedimented at 33 S, as in other mammalian species, and showed a single component of approximately 8500 bases by gel electrophoresis . cDNA was synthesized from the 33-S RNA by using reverse transcriptase in the presence of human placenta ribonuclease inhibitor and in conditions allowing the formation of long transcripts . The latter was made double-stranded using reverse transcriptase and blunt-ended with nuclease S1 . After tailing with dCTP and terminal transferase, the double-stranded cDNA was annealed to pBR322 DNA that had been cleaved at the endonuclease PstI site and tailed with dGTP . The resulting plasmids were used to transform Escherichia coli C600 cells and four cloned recombinants were selected . Each plasmid DNA was shown to contain a sequence complementary to human thyroglobulin mRNA by hybridization with a labeled 33-S mRNA, visualization of cDNA . mRNA hybrids by electron microscopy and filter hybridization selection of mRNA directing the synthesis of immunologically related thyroglobulin peptides in the reticulocyte lysate . The four inserted DNA sequences were 1400 - 1800 base pairs long, two of them showing an homologous sequence of 1100 base pairs . Together, the four cloned DNA fragments represented 63% of the 8500 bases of human thyroglobulin mRNA. Cancer Res, 1981 Nov, 41(11 Pt 1), 4382 - 5 Blocking by the carcinogen, L-ethionine, of SOS functions in a tif-1 mutant of Escherichia coli B/r; Wiesner R et al.; In Escherichia coli, DNA damage by carcinogenic agents results in the coordinate expression of a diversity of functions (SOS functions), many of which are thermally inducible without any damage to DNA in a tif-1 mutant . These include prophage induction, filamentous growth, and an error-prone DNA repair activity, which is responsible for ultraviolet-induced mutagenesis . Ethionine causes hepatic carcinoma in rats after prolonged feeding but is not a mutagen in the Ames test . The present study shows that 10 mM ethionine prevents the thermal induction of lambda-prophage in a tif-1 derivative of E . coli . The enhancement of mutation, which normally occurs at high temperature after a low dose of ultraviolet light, is also blocked by ethionine . Ethionine does not block, to any appreciable extent, the incorporation of radioactive precursors into RNA, DNA, or protein. J Pharm Pharmacol, 1981 Nov, 33(11), 685 - 91 A comparative study on the performance of asbestos-free depth-filters for removal of pyrogens from infusion fluids; Baggerman C et al.; Three asbestos-free depth-filters were compared with traditionally used asbestos-containing depth-filters for chemical integrity, physical integrity and pyrogen retention . With chemical and physical integrity only minor differences between the various filter types were seen . One asbestos-free depth-filter, based on charcoal, showed poor pyrogen-retention; this might be due to a high flow rate used . Asbestos-free depth-filters based on kieselguhr or on a mixture of kaolin and alumina proved to be good alternatives; these filters have proved to be suitable for the removal of pyrogens from electrolyte- or carbohydrate-containing infusion fluids. Nature, 1981 Oct 29, 293(5835), 717 - 22 Expression of a human gene for interferon in yeast; Hitzeman RA et al.; A DNA sequence coding for mature human leukocyte interferon D (LeIF-D) was linked with DNA fragments of the 5'-flanking sequences of the Saccharomyces cerevisiae (yeast) alcohol dehydrogenase I gene in a plasmid capable of autonomous replication and selection in both yeast and Escherichia coli . Yeast cells transformed by these plasmids synthesize up to 1 x 10(6) molecules of biologically active LeIF-D per cell. Biochim Biophys Acta, 1981 Oct 27, 655(3), 374 - 82 Influence of the ionic environment on the in vitro transcription of the spinach plastid DNA by a selectively bound RNA-polymerase DNA complex; Blanc M et al.; The in vitro transcription of chloroplast DNA (ctDNA) is studied using a DNA-protein complex isolated from spinach plastids . The RNA products are compared to the in vivo synthesized ctRNA by competition for hybridization . At least 80% of the in vitro RNA sequences are present in vivo . Modifications of ionic strength or introduction of heparin in the reaction medium has an important effect on transcriptional activity of the complex . Furthermore, the length of the RNA chains increases ionic strength and amount of heparin . The RNA products are analysed by Southern hybridizations to EcoRI cTDNA fragments . Changes in the ionic strength or in the amount of heparin modify heterogeneously the transcription of the various DNA regions . The quantitative distribution of transcripts among the ctDNA fragments is used as evidence for the selectivity of the transcription . The activity of the DNA-protein complex is much more resistant to high ionic strength than an heterologous transcription system using Escherichia coli RNA polymerase and ctDNA . This latter system transcribes less ctDNA fragments. Biochemistry, 1981 Oct 27, 20(22), 6480 - 4 Physical characteristics of the reconstitution intermediates (RI30 and RI30*) from the 30S ribosomal subunit of Escherichia coli; Tam MF et al.; The isolated reconstitution intermediates (RI30 and RI30*) from the 30S ribosomal subunit of Escherichia coli were found to contain ten proteins . The sedimentation coefficients, diffusion coefficients, density increments, extinction coefficients, and molecular weights were determined for the reconstitution particles and compared with those obtained from the 16S rRNA under identical buffer conditions . The results show that the binding of the proteins on the 16S rRNA at 4 degrees C does not markedly affect the folding of the RNA molecule . However, upon heating the RI30 particle at 40 degrees C to form the RI30* particle, significant folding of the RNA took place, giving a structure considerably more compact than that of the 16S rRNA or the RI30 particle. Biochemistry, 1981 Oct 27, 20(22), 6391 - 5 Circular dichroism studies on the interaction of tryptophan synthase with pyridoxal 5'-phosphate; Balk H et al.; The interaction between the beta 2 subunit of tryptophan synthase and the coenzyme pyridoxal 5'-phosphate (PLP) is characterized by induced circular dichroism (CD) in the near-UV (260-285 nm) and in the visible region (320-480 nm, extrinsic Cotton effect) . Because of its high mean residue ellipticity ({theta} = 56 deg cm2 dmol-1 for the isolated holo-beta 2 subunit and 102 deg cm2 dmol-1 for the alpha 2-holo-beta 2 complex, respectively) the latter has been used to define different conformational states of the beta 2 dimer via CD titrations . Fitting the obtained binding parameters to the known data from equilibrium dialysis leads to the result that the low-affinity state of the isolated beta 2 subunit shows a 3 times greater rotational strength than the holoenzyme in the high-affinity state . The generation of the final CD amplitude is characterized by a rate constant intermediate between the values for the formation of the internal aldimine and for the regain of enzymatic activity . Interaction of the alpha 2-apo-beta 2 bienzyme complex with the cofactor leads to a hyperbolic binding curve which is apparently free of contributions caused by unspecific PLP binding outside the active center . The determined dissociation constant of 9 x 10(-7) M is in good agreement with the value of 1 x 10(-6) M as obtained by equilibrium dialysis . Binding kinetics reveal a very slow process with a rate constant of 2.6 x 10(-4) s-1, significantly smaller than that for the regain of catalytic activity during reconstitution of the enzyme. Biochim Biophys Acta, 1981 Oct 27, 655(3), 278 - 90 A kinetic study on the mechanism of inhibition of RNA synthesis catalyzed by DNA-dependent RNA polymerase . Differences in inhibition by ethidium bromide, 3,8-diamino-6-ethylphenanthridinium bromide and actinomycin d; Aktipis S et al.; The mechanism of inhibition of RNA polymerase-catalyzed synthesis of RNA by actinomycin D and the phenan-thridinium derivatives ethidium bromide and 3,8-diamino-6-ethylphenanthridinium bromide (DEMB) is examined . A general kinetic equation describing the dependence of RNA synthesis on DNA template concentration is derived and distinct expressions corresponding to various possible mechanisms of inhibition are subsequently obtained by introducing into the equations assumptions as appropriate for the individual mechanisms . The fitting of the experimental results of inhibition into the resulting equations suggested that the ethidium bromide and DEMB inhibit RNA polymerase by forming an inhibitor-template complex which interferes with enzyme recognition of, and binding to, appropriate sites on the template (binding inhibition) . The fitting of the dependence of the rate of RNA synthesis on the bound-inhibitor to DNA ratios to the derived kinetic expressions also allows a tentative distinction to be made as to whether ethidium bromide and DEMB interfere with RNA synthesis by a mechanism of 'partial' or 'complete' inhibition. Biochim Biophys Acta, 1981 Oct 27, 655(3), 366 - 73 The region of transcriptional initiation in Lytechinus variegatus rRNA genes; Bieber D et al.; A sea urchin ribosomal DNA 1.9 kilobase BamHI fragment adjacent to the 5' end of the 18 S gene has been mapped with the restriction enzymes, XhoI, EcoRi, SmaI and HinfI . A 270 basepair fragment which most likely contains the 5' end of the presumed primary transcript of rRNA was identified by hybridization of {32P}DNA fragments to total nuclear RNA separated on methylmercury hydroxide gels and bound to diazobenzyloxymethyl paper . Under these denaturing conditions the size of Lytechinus variegatus precursor rRNA was determined to be 7.2 kilobases (33 S). J Biol Chem, 1981 Oct 25, 256(20), 10415 - 9 Effect of recA protein on the DNAse activities of the recBC enzyme; Prell A et al.; In Escherichia coli, the recBC enzyme is required for several cellular functions including recombination proficiency, UV resistance, and DNA breakdown following radiation damage to the chromosome, all of which appear to be also under the control of the recA gene . We have studied the influence of purified recA protein on the various nucleolytic activities of the recBC enzyme . Conditions were chosen (with GTP as nucleoside triphosphate) under which recA protein binds to single-stranded DNA without catalyzing D-loop formation and which are favorable for the DNase activities of the recBC enzyme . We found that the degradation of linear duplex DNA was unaffected, but that the endonuclease and exonuclease activities for single-stranded DNA were inhibited by about 50% and 35%, respectively . In contrast, no protection of circular duplex DNA containing single-stranded regions was observed . The results suggest that the recA protein by itself may not act as a potent inhibitor of recBC enzyme-dependent DNA degradation in vivo. J Biol Chem, 1981 Oct 25, 256(20), 10671 - 83 Molecular mechanisms of substitution mutagenesis . An experimental test of the Watson-Crick and topal-fresco models of base mispairings; Sinha NK et al.; The proteins coded by bacteriophate T4 replication genes 32, 41, 43, 44, 45, 61, and 62 together can replicate phi X174 DNA templates very efficiently . The fidelity of this in vitro replication reaction has been measured using an infectivity assay . The product molecules have the same specific infectivity as the template DNA . When an amber mutant DNA template is used, no increase in the frequency of revertants is seen even after more than 60 duplications in vitro . By using imbalances in the concentrations of deoxynucleotide substrates, the error rate during DNA replication in vitro can be greatly increased . Control experiments indicate that the increased mutagenesis is not due to the presence of dITP or dUTP as contaminants in the deoxynucleotide substrates used . The increase in the frequency of revertants is linearly related to the ratio of the correct and the incorrect deoxynucleotides . Determination of the DNA sequence of the revertants induced shows that a change in DNA sequence of the amber site predicted from the nucleotide bias occurs . DNA synthesis in vitro resembles in vivo replication in that the error rate depends not only upon the base change required for reversion but also upon the neighboring DNA sequences . The error rate is estimated to be 5 X 10(-6) at am3 site, 6.4 X 10(-7) at am86 site, and less than 2.9 X 10(-7) at am9 site . Comparison of the frequency of G-T and A-C mispairs reveals that most AT leads to GC transition mutations occur through G-T mispairs . Measurement of the frequency of the mispairs required to induce transversion mutations reveals that these occur primarily through purine-purine mispairs . Transition mutations are more frequent than transversion mutations at both the am3 and the am86 sites . These observations support the models for base pairing errors proposed by Watson and Crick ((1953) Nature 171, 964-967) and Topal and Fresco ((1976) Nature 263, 285-289). J Biol Chem, 1981 Oct 25, 256(20), 10640 - 4 Expression of the Streptomyces enzyme endoglycosidase H in Escherichia coli; Robbins PW et al.; Endoglycosidase H is one of a large number of enzymes secreted by Streptomyces plicatus and other Streptomyces species . When the structural gene for this enzyme is introduced into Escherichia coli attached to the plasmid pBR-322 or Charon 4 phage, the enzyme is synthesized and is found in the periplasmic space, culture medium, and cells . Attachment of the UV-5 lac promoter to a site in the plasmid adjacent to the Streptomyces insert stimulates enzyme synthesis as much as 100-fold . This result demonstrates that transcription of the Streptomyces gene can be initiated from sequences outside of the Streptomyces insert . Initiation of transcription on a Streptomyces promoter is also a suggested but unproven possibility . In contrast to the situation in Streptomyces, where the enzyme has a molecular weight of 27,000, the enzyme made in E . coli has a molecular weight of approximately 30,000 . Possible explanations for this difference in size are lack of cleavage of the Streptomyces secretion "signal sequence" in E . coli or protein "processing" by enzymes secreted into the medium by STreptomyces. Nucleic Acids Res, 1981 Oct 24, 9(20), 5493 - 504 Studies on the selectivity of DNA precipitation by spermine; Hoopes BC et al.; We have examined the selectivity of the precipitation of DNA by spermine . We have found that the intra- and intermolecular condensation of DNA induced by spermine is highly selective even in the presence of added protein or triphosphates . We have also investigated the influence of buffer components on the threshold concentration of spermine required for DNA precipitation . Representative applications exploiting the selectivity of the precipitation reaction are also described. Nucleic Acids Res, 1981 Oct 24, 9(20), 5469 - 82 Determination of DNA cooperativity factor; Amirikyan BR et al.; The paper presents measurements of the difference in the melting temperature of a colE1 DNA region when it is located inside the DNA helix and at its end . A direct comparison of calculations based on the rigorous theory of helix-coil transition with experimental data for .2 M Na+ (the conditions for fully reversible melting) yielded the value of 2.5-5x10(-5) for the cooperatively factor sigma . We discuss the reversibility of DNA melting and the possibility of applying the "all-or-nothing" concept to the melting of DNA regions. Nucleic Acids Res, 1981 Oct 24, 9(20), 5399 - 406 Sequence of a putative promoter region for the rRNA genes of tobacco chloroplast DNA; Tohdoh N et al.; The nucleotide sequence of the segment of tobacco chloroplast DNA adjacent to and including the start of the 16S rRNA gene has been determined . The region just preceding this gene was found to contain a tRNAVal gene and promoter-type sequences similar to those which occur in E . coli were found before this tRNA gene . E . coli RNA polymerase can recognize these sequences and in vitro co-transcribes the tRNA and rRNA genes. Nucleic Acids Res, 1981 Oct 24, 9(20), 5287 - 96 The atp operon: nucleotide sequence of the genes for the gamma, beta, and epsilon subunits of Escherichia coli ATP synthase; Saraste M et al.; The nucleotide sequence of the promoter distal region of the atp (or unc) operon of Escherichia coli has been determined . It encodes the gamma, beta and epsilon subunits of the ATP-synthase complex and includes a noncoding sequence in which transcription of the operon probably terminates . This work completes the nucleotide sequence of the operon which contains nine genes: eight encode structural proteins of the ATP-synthase complex; a ninth, the first in the operon, may be a pilot for assembly . The genes for the alpha and beta subunits have evolved from a common ancestor. Nucleic Acids Res, 1981 Oct 24, 9(20), 5215 - 31 Rifampicin-resistant initiation of DNA synthesis on the isolated strands of ColE plasmid DNA; Boldicke TW et al.; The opposite strands of the ColE1 and ColE3 plasmids were isolated as circular single-stranded DNA molecules . These molecules were compared with M13 and phi X174 viral DNA with respect to their capacity to function as templates for in vitro DNA synthesis by a replication enzyme fraction from Escherichia coli . It was found for both ColE plasmids that the conversion of H as well as L strands to duplex DNA molecules closely resembles phi X174 complementary strand synthesis and occurs by a rifampicin-resistant priming mechanism involving the dnaB, dnaC, and dnaG gene products . Restriction analysis of partially double-stranded intermediates indicates that preferred start sites for DNA synthesis are present on both strands of the ColE1 HaeII-C fragment . Inspection of the nucleotide sequence of this region reveals structural similarities with the origin of phi X174 complementary strand synthesis . We propose that the rifampicin-resistant initiation site (rri) in the ColE1 L strand is required for the priming of discontinuous lagging strand synthesis during vegetative replication and that the rri site in the H strand is involved in the initiation of L strand synthesis during conjugative transfer. Nucleic Acids Res, 1981 Oct 24, 9(20), 5187 - 202 Expression of plant tumor-specific proteins in minicells of Escherichia coli: a fusion protein of lysopine dehydrogenase with chloramphenicol acetyltransferase; Schroder J et al.; Fragment EcoRI 7 from Ti-plasmid pTi Ach5, a part of the T-DNA in octopine tumors, was cloned in both orientations into pACYC184 and expressed in E . coli minicells . The cells synthesized four proteins from four different coding regions on EcoRI 7 . Two of the proteins (Mr 25.000 and 26.000) were expressed with promoters from the Ti-plasmid fragment, while transcription for the two other proteins (Mr 18.000 and 74.000) started with a promoter on pACYC184 . The Mr 18.000 protein represented a fusion product between chloramphenicol acetyltransferase (CAT) on pACYC184 and a part of lysopine dehydrogenase (LpDH), the enzyme synthesizing octopine and lysopine in plant tumor cells . The results suggest that E . coli minicells are a valuable system to study the proteins coded for by the T-region of Ti-plasmids. Nature, 1981 Oct 22, 293(5834), 675 - 7 Steps of mRNA translocation in protein biosynthesis; Holschuh K et al.; The translocation of the messenger RNA relative to the ribosome during peptide synthesis represents an example of a mechano-chemical reaction in which the chemical bond energy of GTP is transformed into coordinated motion . Such transformations also occur during the beating of cilia and flagellae, the contraction of muscle and the migration of chromosomes in cell division . In protein synthesis the functional geometric and energetic conditions for this transformation are well defined . For each peptide bond formed, the ribosome moves one codon along the mRNA (towards the 3' end) and one molecule of GTP is hydrolysed . Although the basic requirements of this reaction have been elucidated, the mechanism is still unresolved . We demonstrate here that translocation can be analysed as a series of binding equilibria shifted by one irreversible, GTP-consuming step . The shift in the binding equilibrium is induced by the transfer of the peptidyl moiety to the (A) site-bound aminoacyl (AA)-tRNA . This results in the A site-bound tRNA having an increased affinity for the high-affinity (P) site, and a strengthened association with the mRNA . Elongation factor (EF) G . GPT catalyses removal of the deacylated tRNA, empties the P site and at the same time loosens ribosome-mRNA association . The result of these changes is that peptidyl(PP)-tRNA . mRNA is shifted from the A site to the P site, binding of AA-tRNA . EF-Tu . GPT to the vacant A site ensuring that the process is irreversible. Biochim Biophys Acta, 1981 Oct 20, 648(1), 71 - 9 Endotoxin-induced alterations in glucose transport in isolated adipocytes; Leach GJ et al.; 2-Deoxyglucose and 3-O-methylglucose were used to assess endotoxin-induced changes in glucose transport in rat adipocytes . 6 h after Escherichia coli endotoxin injection insulin-stimulated 2-deoxyglucose uptake was significantly depressed (V decreased, Km unaltered), phosphorylation of 2-deoxyglucose was seemingly unimpaired; basal 3-methylglucose entry was significantly increased, insulin-stimulated uptake was unaltered . Insulin significantly reduced Km in control and endotoxin-treated cells . Cytochalasin B-insensitive uptake of both 2-deoxy-glucose and 3-methylglucose, a small fraction of total transport, increased significantly in endotoxic cells . Endotoxin reduced spermine- and insulin-stimulated 2-deoxyglucose uptake to a similar extent . Results are consistent with the hypotheses that (1) a site of endotoxin-induced insulin resistance is at the cell membrane level and may reflect a decrease in number of activity of effective carrier units, rather than alterations in affinity, (2) endotoxin does not compromise the hexokinase system, (3) the cell membrane-localized effect of endotoxin on hexose transport is not necessarily mediated by the insulin receptor and (4) the entry of 2-deoxyglucose and 3-methyl-glucose may involve two separate transport systems. Int J Cancer, 1981 Oct 15, 28(4), 485 - 96 Expression of SV40 T antigen polypeptides in cells biochemically transformed by plasmids containing the herpes simplex virus thymidine kinase gene and the genome of an SV40tsA mutant; Kit S et al.; To study the expression of SV40 tsA genomes that had been non-selectively introduced into mouse cells, SV40 tsA207 DNA was cleaved with BamH I and ligated to BamH I-cleaved plasmid pAGO DNA, which contains a functional HSV-1 thymidine kinase (TK) gene in the form of 2 kbp Pvu II fragment inserted at the Pvu II site of pBR322 . Recombinant plasmids (11-12 kbp) were isolated and amplified in E . coli K12 strain RRI . Restriction nuclease analyses demonstrated that recombinant plasmids pSB15 and pSB10 contained intact SV40 genomes with the polarity of transcription oriented in the same direction (clockwise) or the opposite direction (counterclockwise), respectively, in relation to that of the HSV-1 TK gene . Cla I-cleaved pSB10 and pSB15 DNAs were used to transform LM(TK-) cells to TK+ . Serological and disc PAGE analyses showed that clonal lines transformed by these plasmids all expressed the selected marker, HSV-1 TK . Molecular hybridization experiments showed that transformed clonal lines TF pSB10 C7 and TF pSB15 C10 had integrated intact SV40 genomes at one integration site, TF pSB10 C3 had integrated an SV40 genome with a small deletion near the BamH I site, but TF pSB15 Cl had integrated a plasmid from which most of the SV40 nucleotide sequences had been deleted . IF assays with hamster anti-SV40 tumor sera showed that TF pSB10 C7 and TF pSB15 C10 strongly expressed SV40 T antigens in over 90% of the cells, TF pSB10 C3 expressed SV40 T antigens in a minority of the cells, and TF pSB15 C1 did not express SV40 T antigens at all . {35S}-methionine labelling and immunoprecipitation experiments showed that, at 36.5 degrees C: (1) TF pSB10 C7 and TF pSB15 C10 expressed 92K and 20K mol . wt . species of SV40 T antigens and 50-55K cellular protein; (2) expression of all three was reduced in TF pSB10 C3 cells; and (3) TF pSB15 C1 expressed none of the SV40 T antigens, nor did parental LM(TK-) or TF 8-2 transformed cells (which contained the HSV-1 TK gene but not SV40 DNA) . At 40 degrees C, labelling of the 50-55K cellular protein was markedly reduced in TF pSB10 C7 and pSB15 C10 cells . The results suggest that SV40 large T antigen (92K) induces and/or stabilizes the 50-55K cellular protein in these mouse cells. Biochemistry, 1981 Oct 13, 20(21), 6235 - 44 Nonrandom substitution of 2-aminopurine for adenine during deoxyribonucleic acid synthesis in vitro; Pless RC et al.; The incorporation of the deoxyribonucleotide of 2-aminopurine {(AP)} for deoxyadenylate into deoxyribonucleic acid (DNA) in vitro has been examined by using five highly purified DNA polymerases: calf thymus polymerase alpha, Escherichia coli polymerase I, and the polymerases induced by T4 phage mutant L56 (mutator phenotype), wild-type T4 phage, and T4 phage mutant L141 (antimutator phenotype) . On a template of gapped salmon sperm DNA, the overall incorporation of (AP) relative to the incorporation of adenine decreases in this series of enzymes, in line with the increasing 3'-exonucleolytic activity associated with these polymerases . The nearest-neighbor distributions for (AP) and for adenine in the newly synthesized DNA were determined to test for potential sequence selectivity in the incorporation of (AP) . In polymerizations in which d(AP)TP fully replaced dATP, the L141 polymerase, and to a lesser degree the wild type T4 polymerase, synthesized a DNA in which the distribution for (AP) was distinctly skewed compared to the nearest-neighbor distribution observed for adenine; incorporation of (AP) was relatively favored after guanine and disfavored after adenine and thymine . These sequence effects were less pronounced in syntheses in which both dATP and d(AP)TP were present . When dGTP was replaced by dITP, or dTTP by dUTP, adenine was still incorporated to the normal extent after the analogue, but the incorporation of (AP) was reduced after these analogues, which form weaker base pairs . The results indicate that incorporation of (AP) is disfavored with all polymerases tested and that a greater bias exists with those polymerases containing an active 3'-exonuclease . This bias against (AP) incorporation is alleviated after strong base pairs, and particularly following guanine, possibly due to stabilizing vertical stacking interactions. Biochemistry, 1981 Oct 13, 20(21), 6164 - 9 Unfolding of lac repressor and its proteolytic fragment by urea: headpieces stabilize the core within lac repressor; Schnarr M et al.; Circular dichroism measurements were used to compared the urea-induced unfolding transition of the lac repressor with those of its separated tryptic fragments, the tetrameric core, and the N-terminal headpiece . The presence of the headpieces covalently linked to the core in the intact repressor leads to a stabilization against urea denaturation as compared to that for the isolated core . This results in a shift of the midpoint of the transition by about 0.5 M urea . When the inducer isopropyl beta-D-thiogalactoside is bound, the core is stabilized more than the entire repressor . The isolated headpiece is considerably more stable against urea denaturation than the tryptic core or the lac repressor . The reversible denaturation process of the headpiece was quantitatively analyzed, and the free energy of unfolding in the absence of urea was found to be 2.4 or 2.9 kcal/mol, depending on the method of calculation used . Comparison between the circular dichroism spectra of the lac repressor, the tryptic core of the lac repressor, and the headpiece supply further evidence that there are no major conformational differences between the structural domains (core and headpieces) before and after proteolytic cleavage of the lac repressor . These results are discussed with respect to the contacts between the different domains of the protein . It is concluded that relatively weak interdomain contacts are probably responsible for the stabilization of the core by the covalently linked headpieces and that these contacts might be weakened upon binding of the inducer. Biochemistry, 1981 Oct 13, 20(21), 6109 - 18 lac Repressor: a proton magnetic resonance look at the deoxyribonucleic acid binding fragment; Arndt KT et al.; The DNA binding fragment from Escherichia coli lac repressor, the N-terminal 56 amino acid residue "headpiece", has been examined by high-resolution 1H NMR spectroscopy at 360 MHz . The aromatic region has been examined in detail along with the four headpieces of altered repressors that are each missing one of the tyrosines, respectively . The spectra here show more resolved resonances and correct errors in the resonance assignments that have been published by Ribeiro et al . (1981b) Ribeiro, A . A., Wemmer, D., Bray, R . P., Wade-Jardetzky, N . G., & Jardetzky, O . (1981) Biochemistry 20, 818-823} . These corrections allow an interpretation of the spectroscopic observations that is now consistent with the extensive genetic analysis that has been carried out with the lac repressor gene . In addition, nuclear Overhauser enhancement measurements give a guide to the interresidue distances among the aromatic residues in this protein fragment. Biochemistry, 1981 Oct 13, 20(21), 6103 - 8 Synthesis of a nitrobenzeneboronic acid substituted polyacrylamide and its use in purifying isoaccepting transfer ribonucleic acids; Johnson BJ; Highly purified isoaccepting species of transfer ribonucleic acid (tRNA) were prepared by use of a polyacrylamide substituted with nitrobenzeneboronic acid functional groups . This method exploits the well-known ability of boronic acids to complex with RNA cis-diols . tRNA isoacceptors were obtained by enzymatically acylating a mixture of tRNA species with a single amino acid and passing the mixture over a solid-state nitrobenzeneboronic acid at pH 6.5 or 7.0 . Pure aminoacyl-tRNA eluted at the column liquid volume, and unacylated tRNA species were bound . The bound species were recovered by lowering the pH of the eluant to 4.5 . This procedure is uncomplicated, rapid, and applicable to nearly all tRNA isoacceptors . It does not chemically modify the tRNA(s) of interest or adversely affect their ability to be aminoacylated . Since boronic acids must be ionized to complex with cis-1,2-diols, boronic acid derivatives were prepared which ionize at a pH compatible with the stability of the aminoacyl bond . Two isomeric benzeneboronic acids with pKas of 6.8 and 7.4 were synthesized by introducing electron-withdrawing nitro groups into the aromatic ring . The addition of succinyl side chains permitted the nitrobenzeneboronic acids to be coupled to aminoethylpolyacrylamide . The properties of the nitrobenzeneboronic acid substituted acrylamide were illustrated by enriching phenylalanyl-tRNA at pH 7.0 to greater than 95% purity (1.63 nmol of phenylalanine accepted per A260 unit of tRNA) and seryl-tRNA isoacceptors at pH 6.5 to essentially theoretical purity (1.58 nmol of serine accepted per A260 unit of tRNA. Biochemistry, 1981 Oct 13, 20(21), 6018 - 23 Reversible inactivation of Escherichia coli methionyl-tRNA synthetase by covalent attachment of formylmethionine tRNA to the tRNA binding site with a cleavable cross-linker; Schulman LH et al.; Protein affinity labeling groups have been attached to single-stranded cytidine residues in four structural regions of tRNAfMet . Modification of the tRNA with an average of one cross-linking group per molecule is achieved with retention of 75% of the original methionine acceptor activity . Incubation of the modified tRNA with methionyl-tRNA synthetase (MetRS) results in covalent coupling of the protein and nucleic acid by reaction of N-hydroxysuccinimide ester groups attached to the tRNA with lysine residues in the enzyme . In the presence of excess MetRS, approximately 30% of the input tRNA can be covalently bound to protein, indicating that lysine residues are appropriately oriented for reaction with cross-linking groups attached to certain sites in the tRNA but not to others . The cross-linking reaction results in loss of aminoacylation activity of MetRS equal to the amount of covalently bound tRNA . Enzyme activity is restored by release of bound tRNA following cleavage of the disulfide bond of the cross-linker with a sulfhydryl reagent . The data indicate that cross-linking occurs at the tRNA binding site of the enzyme . In the presence of excess modified tRNAfMet, a maximum of 1 mol of tRNA is cross-linked per mol of MetRS, in keeping with the known anticooperative tRNA binding properties of the native dimeric synthetase . In addition, the coupling reaction is effectively inhibited by unmodified tRNAfMet, but not by noncognate tRNAs. Biochim Biophys Acta, 1981 Oct 13, 661(2), 261 - 6 Studies on aspartase . VII . Subunit arrangement of Escherichia coli aspartase; Watanabe Y et al.; Aspartase (L-aspartate ammonia-lyase, EC 4.3.1.1) of Escherichia coli is composed of four subunits of seemingly identical molecular weight (Suzuki, S., Yamaguchi, J . And Tokushige, M.(1973) Biochim . Biophys . Acta 321, 369-381) . The subunit arrangement of the enzyme was studied by two distinct methods, cross-linking of subunits with a bifunctional reagent, dimethyl suberimidate, and statistical classification of negatively stained electron microscopic images . In the former method, the densitometric patterns of the cross-linked aspartase were analyzed quantitatively after separating each component by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and the results were compared with the theoretical distribution . In the latter method, a number of electron microscopic images were classified into several groups according to their characteristic appearance . The results obtained by these two methods are compatible with the possibility that the enzyme has a tetrameric structure consisting of two pairs of dimers, in which the two pairs of rod-shape subunits meet perpendicularly, being typical of D2 symmetry. Biochemistry, 1981 Oct 13, 20(21), 6272 - 9 Reconstitution of Escherichia coli membrane vesicles with D-amino acid dehydrogenase; Olsiewski PJ et al.; When purified D-amino acid dehydrogenase {Olsiewski, P . J., Kaczorowski, G . J., & Walsh, C . T . (1980) J . Biol . Chem . 255, 4487} is incubated with right-side-out membrane vesicles from Escherichia coli, the enzyme binds to the membrane in a time- and concentration-dependent manner . As a result, the vesicles acquire the ability to oxidize D-alanine and catalyze D-alanine-dependent active transport . Similarly, incubation of D-amino acid dehydrogenase with inside-out vesicles results in binding of enzyme and D-alanine oxidase activity . Antibody inhibition studies indicate that the enzyme is bound exclusively to the inner cytoplasmic surface of the membrane in native vesicles (i.e., membrane vesicles prepared from cells induced for D-amino acid dehydrogenase) . In contrast, similar studies with reconstituted vesicles demonstrate that enzyme binds to the surface exposed to the medium regardless of the orientation of the membrane . Thus, enzyme bound to right-side-out vesicles is located on the opposite side of the membrane from where it is normally found . Remarkably, in the presence of D-alanine, reconstituted right-side-out and inside-out vesicles generate electrochemical proton gradients of similar magnitude but opposite polarity, indicating that enzyme bound to either surface of the membrane is physiologically functional . The results suggest that vectorial proton translocation via the respiratory chain occurs at a point distal to the site where electrons enter the respiratory chain from the primary dehydrogenase, a conclusion that is inconsistent with the notion that the dehydrogenase forms part of a proton-translocating loop. Nucleic Acids Res, 1981 Oct 10, 9(19), 5125 - 40 Mapping tRNA structure in solution using double-strand-specific ribonuclease V1 from cobra venom; Lockard RE et al.; A method for mapping all base-paired stems in both elongation and initiator tRNAs is described using double-stranded-specific ribonuclease V1 from the venom of the cobra Naja naja oxiana . 32p-end-labeled RNA is first partially digested with double-strand-specific V1 nuclease under near physiological conditions, and the resultant fragments are than electrophoretically fractionated by size in adjacent lanes of a polyacrylamide gel run in 90% formamide . After autoradiography, the base-paired nucleotides are definitively located by comparing V1 generated bands with fragments of known length produced by both Neurospora endonuclease and base-specific ribonucleases . Using the substrates yeast tRNAPhe an E, coli tRNAfMet of known three-dimensional structure, we find V1 nuclease to cleave entirely within every base-paired stem . Our studies also reveal that nuclease V1 will digest paired nucleotides not hydrogen-bonded by standard Watson-Crick base-pairing . In yeast tRNAPhe cleavage of both wobble base-pairs and nucleotides involved in tertiary base-base hydrogen bonding is demonstrated. J Biol Chem, 1981 Oct 10, 256(19), 9778 - 81 Free sigma factor of Escherichia coli RNA polymerase can bind to DNA; Kudo T et al.; Free sigma factor from E . coli RNA polymerase holoenzyme was shown to associate with supercoiled pBR350 and pBRH4 plasmid DNA s by two methods . The banding pattern of sigma factor through a nondenaturing polyacrylamide slab gel was significantly altered in the presence of supercoiled DNA; sigma factor had little or no affinity to linear, double- or single-stranded DNA . At saturation, approximately one sigma factor was bound/200 base pairs of supercoiled pBR350 DNA . By the nitrocellulose filter trapping method, sigma factor was able to bind supercoiled pBRH4 DNA much more efficiently than linear double-stranded pBRH4 DNA . These results suggest that sigma factor plays a role in the binding of holoenzyme to DNA and may be involved in locally denaturing DNA in the promoter region as postulated by previous investigators. J Biol Chem, 1981 Oct 10, 256(19), 9770 - 3 Mistranslated protein in Escherichia coli; Parker J; Amino acid starvation of a variety of different types of cells has been reported to induce protein degradation and also specific mistranslation . For certain amino acid starvations, the mistranslated protein, which contains specific amino acid substitutions, can be separated and quantified by two-dimensional polyacrylamide gel electrophoresis . In this paper, I show that this specifically mistranslated protein, made during amino acid starvation, does not seem to be preferentially degraded during continued starvation or renewed growth . Specifically mistranslated ribosomal protein is also assembled into ribosomes in the same proportion that it is made . These results imply that the amino acid substitutions apparently made (lysine for asparagine or glutamine or histidine) do not lead to proteins recognized as grossly "abnormal" by the cell's proteolysis systems. J Biol Chem, 1981 Oct 10, 256(19), 9959 - 65 The anaerobic sn-glycerol-3-phosphate dehydrogenase of Escherichia coli . Purification and characterization; Schryvers A et al.; We have purified the membrane-extrinsic anaerobic glycerol-3-phosphate dehydrogenase from Escherichia coli using a strain harboring a recombinant ColE1:E . coli plasmid carrying the glpA gene . The purified enzyme is composed of subunits of 62,000 and 43,000 molecular weight in 1:1 molar ratio as determined by medium dodecyl sulfate polyacrylamide gel electrophoresis, sedimentation equilibrium, and chemical cross-linking studies . The presence of 20% ethylene glycol stabilizes the enzyme by preventing dissociation of the subunits . The purified enzyme contains 1 mol of noncovalently bound FAD and 2 mol of non-heme iron/dimer . The FAD can be reduced by addition of substrate and resides in the large subunit . Addition of exogenous flavins stimulates the rate of the enzymatic reaction, and the effects of FAD and FMN can be distinguished by the following properties: (i) FAD causes a 20% increase in enzymatic activity with a half-maximal concentration of 200 nM whereas FMN results in a 6-fold increase in activity with a half-maximal concentration of 130 microM . (ii) When methylene blue replaces phenazine methosulfate as an oxidation-reduction coupling dye, FAD stimulates the rate of the reaction, whereas FMN inhibits it . Limited proteolysis or treatment with sulfhydryl reagents does not affect activity but removes capacity for stimulation by exogenous flavins. J Biol Chem, 1981 Oct 10, 256(19), 9883 - 9 On the fidelity of DNA replication . Effect of the next nucleotide on proofreading; Kunkel TA et al.; The contribution of proofreading to the fidelity by which Escherichia coli DNA polymerase I copies natural DNA has been analyzed by two independent criteria . With phi X174 am 3 DNA as a template, there is approximately a 25-fold increase in noncomplementary base substitutions at position 587 when the concentration of the next correct nucleotide, dATP, is increased . Sequence analysis indicates that the mistakes represent misincorporation of C in place of T at position 587 . This mutagenic response is presumed to result from a decrease in the probability of excision by the 3' leads to 5' exonuclease of Pol I and is considered within the context of current theories on proofreading . No enhanced mutagenicity is observed with avian myeloblastosis virus DNA polymerase, which lacks a 3' leads to 5' exonuclease . Using a second approach, an enhancement in mutagenesis as large as 30-fold is observed to result from the addition of deoxynucleoside monophosphates to the Pol I reaction . This mutagenicity occurs with any of the four deoxynucleoside monophosphates and is independent of a significant inhibition of DNA synthesis, thus supporting proofreading models in which sites of excision and incorporation are independent . The results of both approaches suggest that the exonucleolytic activity of Pol I can increase fidelity by approximately 30-fold on natural DNA, a value much higher than previous estimates with polynucleotide templates . The effect of the next correct nucleotide in decreasing accuracy provides an in vitro probe for screening eukaryotic cells for putative proofreading functions. J Biol Chem, 1981 Oct 10, 256(19), 10156 - 60 Does simian virus 40 T antigen unwind DNA? Myers RM, Kligman M, Tjian R. Recently developed techniques allow rapid and accurate determination of protein-induced DNA unwinding . We have used such techniques to determine whether SV40 T antigen unwinds DNA . Assays were performed with purified wild type T antigen, D2 protein and the SV80 T protein over a wide range of protein concentrations . Additionally, several DNA templates were used, including SV40 DNA, pBR322 DNA, and two plasmid DNAs containing one and three copies of the SV40 origin of replication . None of the purified proteins showed unwinding activity either in the presence or absence of ATP and Mg2+ . As a positive control, we found that unwinding of pBR322 DNA by Escherichia coli RNA polymerase occurred under identical assay conditions . Protein-DNA interactions that involve unwinding of the helix are sensitive to the superhelical density of the DNA . Using a filter binding assay, we found that the efficiency of the binding of T antigen to the viral origin of replication does not depend upon the topological state of the DNA . This result further supports our conclusion that SV40 T antigen is not an unwinding protein. Nucleic Acids Res, 1981 Oct 10, 9(19), 5109 - 24 Secondary structure formation during RNA synthesis; Kramer FR et al.; We observed the secondary structures that formed in an RNA molecule during its synthesis . Some of the secondary structures seen in nascent chains were observed to form, then to dissociate in favor of an alternative structure, and then to reform, as chain growth continued . The results show that secondary structures in an RNA molecule are in a state of dynamic equilibrium, and that the extension of a sequence by chain growth, or the reduction of a sequence by processing, may result in significant changes in the secondary structures that are present. Exp Hematol, 1981 Oct, 9(9), 950 - 5 Hematopoietic response of splenectomized C3HeB/FeJ and C3H/HeJ mice to lipopolysaccharide; MacVittie TJ et al.; The paired, inbred mouse strains C3HeB/FeJ and C3H/HeJ differ in their extramedullary hemopoietic response to lipopolysaccharide (LPS) . The C3H/HeJ exhibit reduced responses relative to the C3HeB/FeJ in terms of colony-stimulating factor, endogenous and exogenous spleen-derived stem cells (CFUS), and spleen-derived granulocyte-macrophage colony-forming cells (GM-CFC) whereas equivocal responses were noted for the marrow-derived CFUS and GM-CFC . In an effort to determine if the mutational defect was specifically limited to extramedullary expression, we utilized C3HeB/FeJ mice and C3H/HeJ mice at 6 weeks postsplenectomy . The splenectomy emphasizes the medullary response to an i.p . injection of 10 microgram Escherichia coli, LPS-W . Significant differences were revealed in the responses of C3HeB/FeJ and C3H/HeJ marrow-derived cells . Total cells, CFUS, GM-CFC, and the macrophage colony-forming cell (M-CFC) in the C3HeB/FeJ strain all decreased significantly from their saline-injected control values as well as from their counterpart C3H/HeJ values within 48 h after injection of LPS-W . The decline in C3HeB/FeJ CFUS, GM-CFC, and M-CFC was followed by an overshoot, with subsequent return to control values . The total nucleated cells, CFUs, GM-CFC, and M-CFC derived from the C3H/HeJ marrow were characteristically nonresponsive and never varied significantly from their saline-injected control values over the observation period . These results suggest that the phenotypic effect of the defective gene can extend to all hemopoietic tissue, medullary and extramedullary, that normally respond to the factors released through expression of the LPS locus. Pol J Pharmacol Pharm, 1981 Oct, 33(3), 305 - 12 Effects of 5,7-dihydroxytryptamine and 6-hydroxydopamine on fever response in conscious rats; Matuszek M et al.; The effects of 5,7-dihydroxytryptamine (5,7-DHT) and 6-hydroxydopamine (6-OHDA) on the febrile response to E . coli lipopolysaccharide (LPS) of unanesthetized rats examined . Depleting serotonin (5-HT) in brain with 5,7-DHT produced an attenuation in fever response to LPS, while depleting noradrenaline (NA) content in the preoptic area with 6-OHDA produced an opposite effect . However, 6-OHDA when given intraventricularly (icv) was without any significant effect on fever response to LPS . Presented data indicate that alterations in both noradrenergic and serotoninergic system of the rat brain affect the febrile response response to bacterial pyrogen . Moreover, one might conclude that integrity of noradrenergic neurons in central nervous system (CNS) in the rat is not essential for appearance of pyrogen fever. J Biochem (Tokyo), 1981 Oct, 90(4), 1033 - 45 A large-scale preparation and some physicochemical properties of recA protein; Kuramitsu S et al.; Pure recA protein was easily obtained from Escherichia coli harboring plasmid pTM-2 which carried the recA gene by two chromatographic steps on phosphocellulose and DEAE-cellulose . RecA protein was stable in the pH range of 6 to 9 at 25 degrees C . RecA protein was found to aggregate highly under these conditions . Lowering of the protein concentration, the presence of glycerol, and lowering of the pH in the pH stability region diminished the extent of aggregation . The spectroscopic properties of recA protein were measured in the presence of 10% (v/v) glycerol . RecA protein had an absorption maximum at 278 nm . The value of a1% 1cm at 278 nm was determined to be 5.7 . The tryptophyl fluorescence spectrum excited at 295 nm had an emission maximum at 340 nm and the quantum efficiency of recA protein relative to N-acetyl-L-tryptophanamide was determined to be 0.65 . The CD spectrum of recA protein had negative double maxima at 210 and 220 nm . The alpha-helical content was estimated to be about 40% from the CD spectrum in the region of 200 to 250 nm . All three cysteinyl residues of recA protein were reacted with 5,5'-dithiobis(2-nitrobenzoic acid), and recA protein was found to have neither intramolecular nor intermolecular disulfide bond . The reactivities of the SH groups were changed by the presence of ATP or ADP . The denaturation of recA protein by guanidine hydrochloride was studied by measuring CD at 220 nm and tryptophyl fluorescence . The denaturation curve obtained by CD measurement consisted of two stages, one of which lies between 0 and 1.8 M and the other above 1.8 M guanidine hydrochloride . On the other hand, the denaturation curve obtained by fluorescence measurement consisted of a single transition in the concentration range of about 1 to 2.3 M guanidine hydrochloride. J Am Vet Med Assoc, 1981 Oct 1, 179(7), 673 - 6 External cardiovascular resuscitation of the anesthetized pony; Frauenfelder HC et al.; External cardiac massage and concomitant respiratory support were used successfully 6 of 8 anesthetized ponies sustaining unexpected cardiac arrest while being used in a study of shock . Approximately 20 thoracic compressions/min maintained systolic and diastolic aortic blood pressures in excess of 50% of the corresponding base-line values in 5 ponies . The high success rate was attributed to early recognition of the problem, the small size of the patient, and the relatively short duration of cardiopulmonary resuscitation (average, 2.9 minutes) . It was concluded that external cardiac message can be effective for cardiopulmonary resuscitation in selected equine patients that have sustained cardiac arrest. J Gen Microbiol, 1981 Oct, 126(Pt 2), 277 - 87 The reaction of cytochrome o in Escherichia coli K12 with oxygen . Evidence for a spectrally and kinetically distinct cytochrome o in cells from oxygen-limited cultures; Poole RK et al.; Intact cells harvested from O2-limited batch cultures of Escherichi coli K12 contained high levels of the CO-binding cytochromes d, o and a1 . In photodissociation difference spectra (i.e . photolysed minus reduced + CO), a peak at 436 nm and a trough at 415 nm have been assigned to an 0-type cytochrome, and not cytochrome d, by photolysis with white light and an He-Ne laser . The reaction of reduced cytochrome o436 with O2 at sub-zero temperatures involved O2 binding to give intermediate(s) with spectral characteristics similar to those of the reduced oxidase-CO complex . The reaction with O2 at successively higher temperatures (range -98 to -59 degrees C) was accompanied by the formation of a trough (with reference to the CO-liganded state) at 436 nm which eventually shifted to 432 nm, indicative of the oxidized form . The apparent energy of activation at low temperatures was 44.6 kJ mol-1 (10.7 kcal mol-1) . There was a linear relationship between the rate of formation of the oxygen compound and the O2 concentration up to about 0.5 mM . The second-order constant for this reaction was 10.9 M-1 s-1 at 100 degrees C, at least 10-fold greater than for the reaction of cytochrome o432 with O2 in cells from vigorously aerated cultures . The reaction of both types of cytochrome o with O2 was not readily reversible in the light or in the dark and was further distinguished from the reaction with CO by the markedly lower velocity of the CO reaction . Comparisons are drawn between the reactions with O2 of cytochrome(s) o in E . coli from O2-sufficient and O2-limited cultures and of mitochondrial cytochrome a3 . It is proposed that, like the synthesis of cytochrome d, the formation of cytochrome o436 represents an adaptation of the organism to reduced O2 availability. Aust Vet J, 1981 Oct, 57(10), 458 - 60 Colisepticaemia of lambs; Mason RW et al.; In 1979 and 1980, 23 and 29 cases of ovine colisepticaemia from 16 and 18 properties respectively were diagnosed . All cases but one were caused by Escherichia coli 078: NM (Non Motile) . Mortality ranged from 1% to 5% with an age distribution of 3 to 12 weeks . Most cases occurred in October . Thirty-three of the isolates were tested against a wide range of antibiotics and all had similar sensitivity patterns . E . coli 078 was detected less frequently in rectal cultures than in bile, small intestinal or nasal cultures from colisepticaemic lambs . In addition, E . coli 078 was recovered more frequently from nasal than from rectal cultures of lambs dying from causes other than colisepticaemia in colisepticaemic flocks . These findings suggest that perhaps the nasal route may be the more important portal of infection in ovine colisepticaemia caused by E . Coli 078 organisms. Mutat Res, 1981 Oct, 83(3), 307 - 19 Effects of the umuC36 mutation on ultraviolet-radiation-induced base-change and frameshift mutations in Escherichia coli; Kato T et al.; The effects of the umuC36 mutation on the induction of base-change and frameshift mutations were studied . An active umuC gene was necessary in either the uvr+ strains of Escherichia coli K12 for UV- and X-ray-induced mutations to His+, ColER and SpcR, which are presumably base-change mutations, but it was not essential for ethyl methanesulphonate or N-methyl-N'-nitro-N-nitrosoguanidine-induced His+ mutations . In contrast, only 1 out of 13 trp- frameshift mutations examined was UV reversible, and the process of mutagenesis was umuC+-dependent, whereas a potent frameshift mutagen, ICR191, effectively induced Trp+ mutations in most of the strains regardless of the umu+ or umuC genetic background . These results suggest that base substitutions are a major mutational type derived from the umuC+-dependent pathway of error-prone repair. Proc Soc Exp Biol Med, 1981 Oct, 168(1), 33 - 7 Modulation of lymphoid cell blastogenic responsiveness to mitogens by Nippostrongylus brasiliensis infection; Vickery AC et al.; Spleen and mesenteric lymph node cell blastogenic responses to the mitogens concanavalin A and lipopolysaccharide and to parasite antigens were examined in vitro following removal from mice undergoing primary or secondary infection with Nippostrongylus brasiliensis . During primary infection spleen cells showed a marked increase in proliferative responsiveness to both mitogens, followed by a marked depression thereafter . During a secondary infection the response of spleen cells to both mitogens remained depressed . In contrast, cells from the mesenteric lymph nodes of infected mice exhibited enhanced responsiveness to Con A and LPS, followed by depression of the response, followed by another cycle of enhancement upon reinfection . Sensitivity of both spleen and especially mesenteric lymph node cells to Nb antigens was greatest at approximately the time of worm expulsion: Day 13 after primary and Day 8 after secondary infection. J Neurochem, 1981 Oct, 37(4), 998 - 1005 Determination of brain enolase isozymes with an enzyme immunoassay at the level of single neurons; Kato K et al.; Ultrasensitive enzyme immunoassay systems for the assay of rat brain enolase isozymes (alpha alpha, alpha gamma, and gamma gamma forms) were prepared by use of beta-D-galactosidase from Escherichia coli as label and the purified rabbit antibodies to alpha alpha and gamma gamma enolases . The antibodies were purified from the immunoglobulin G (IgG) fractions of antisera by immunoaffinity chromatography with a column of the corresponding antigen-coupled Sepharose . Sandwich-type immunoassay systems with the galactosidase-labeled antibody Fab' fragments and the antibody F(ab')2-immobilized polystyrene beads could determine amounts as small as 1 amol (10(-18) mol) of each isozyme . Purkinje cell bodies picked up from the bulk-separated fraction by means of a nylon loop were subjected to the assay at the level of single cells . In contrast to previous report, this neuron contained not only the gamma gamma but also the alpha gamma and alpha alpha enolases at a level of amol per cell body, although the concentration of gamma gamma was the highest . Immunohistochemical experiments on te cerebellum with the peroxidase-labeled antirabbit IgG antibody and the unlabeled antibody method confirmed the above results, and indicated that both alpha and gamma subunits of hte enolase were stained intensely in axons. J Dairy Res, 1981 Oct, 48(3), 379 - 86 Evidence of penetration of the bovine teat duct by Escherichia coli in the interval between milkings; Bramley AJ et al.; In 3 consecutive experiments, each using 20 cows, the application of Escherichia coli to teat ends after milking led to high rates of intramammary infection . These infections were not prevented by disinfection of the teats before milking, by the installation of shields in the short milk tubes of th milking cluster or by the use of an individual quarter milking cluster . Rates of infection were significantly lower when teat contamination was applied 1 h before milking compared to contamination applied immediately after milking . These data suggest that penetration of te teat duct by the E . coli occurred in the period between contamination and milking . Seventy four percent of infections occurred in hindquarters and there were variations in the susceptibility of cows to infection. Photochem Photobiol, 1981 Oct, 34(4), 529 - 33 Roles of the relA(+) gene and of 4-thiouridine in near-ultraviolet (344 nm) radiation inhibition of induced synthesis of tryptophanase in Escherichia coli B/r; Sharma RC et al.; Near-ultraviolet radiation (near UV; 300-380 nm) is known to inhibit the induced synthesis of tryptophanase by tryptophan in Escherichia coli, showing an action spectrum similar to that for near-UV-induced growth delay . The present work shows that a relA mutant of E . coli B/r exhibits 50% as much monochromatic near-UV (334 nm) inhibition of tryptophanase induction as the wild type, and tht a mutant lacking 4-thiouridine, an unusual nucleoside in tRNA, exhibits greater than 10% as much inhibition of tryptophanase induction . These findings indicate that 4-thiouridine is almost the sole chromophore for this effect in E . coli B/r, but that only 50% of the effect operates by a mechanism utilizing the relA(+) gene product; growth delay appears not to be primarily involved. Photochem Photobiol, 1981 Oct, 34(4), 465 - 9 Pyrimidine dimers induced in Escherichia coli DNA by ultraviolet radiation present in sunlight; Ellison MJ et al.; Escherichia coli DNA was irradiated with various wavelengths of monochromatic UV light from 254 to 320 nm, and the relative yields of the different cyclobutane pyrimidine dimers determined . Cytosine-thymine dimers (C mean value of T) were more frequent than thymine dimers (T mean value of T) at low fluences of 300 and 313 nm light, whereas the reverse was true at either longer or shorter wavelengths . Thus, in the solar UV range deemed responsible for skin cancer (i.e . 295-315 nm), C mean value of T are probably more important than T mean value of T. Proc Natl Acad Sci U S A, 1981 Oct, 78(10), 6344 - 8 Potential for selection among nearly neutral allozymes of 6-phosphogluconate dehydrogenase in Escherichia coli; Hartl DL et al.; Six gnd alleles coding for naturally occurring allozymes of 6-phosphogluconate dehydrogenase {6-phospho-D-gluconate:NAD(P)+ 2-oxidoreductase, EC 1.1.1.43} have been transferred by transduction into the genetic background of Escherichia coli K-12 and examined for their selective effects in chemostats in which gluconate was limiting . Four of the alleles are evidently neutral or nearly neutral, inasmuch as their selective effects, if any, fall below the limit of resolution of the procedure--0.5%/hr or about 1% per generation . One allele is detrimental in limiting gluconate but not in limiting glucose or fructose . Another allele has a detrimental, density-dependent, epistatic interaction with tonA . We suggest that all six alleles are neutral or nearly neutral in natural populations but that they are not functionally equivalent; their functional differences are potentially important because they can become expressed as differences in fitness under the appropriate conditions of environment or genetic background . Under these conditions, otherwise neutral alleles can become subject to selection. Proc Natl Acad Sci U S A, 1981 Oct, 78(10), 6271 - 5 pH homeostasis in Escherichia coli: measurement by 31P nuclear magnetic resonance of methylphosphonate and phosphate; Slonczewski JL et al.; The intracellular pH of Escherichia coli cells, respiring on endogenous energy sources, was monitored continuously by 31P NMR over an extracellular pH range between 5.5 and 9 . pH homeostasis was found to be good over the entire range, with the data conforming to the simple relationship intracellular pH = 7.6 + 0.1(external pH - 7.6) so that the extreme values observed for intracellular pH were 7.4 and 7.8 at external pH 5.5 and 9, respectively . As well as inorganic phosphate, we employed the pH-sensitive NMR probe methylphosphonate, which was taken up by glycerol-grown cells and was nontoxic; its pKa of 7.65 made it an ideal probe for measurement of cytoplasmic pH and alkaline external pH. Proc Natl Acad Sci U S A, 1981 Oct, 78(10), 6061 - 5 tif-1 mutation alters polynucleotide recognition by the recA protein of Escherichia coli; McEntee K et al.; The requirements for polynucleotide-dependent hydrolysis of ATP and for proteolytic cleavage of phage lambda repressor have been examined for both the wild-type (recA+ protein) and the tif-1 mutant form {tif(recA) protein} of the recA gene product . The recA+ and tif(recA) proteins catalyze both reactions in the presence of long single-stranded DNAs or certain deoxyhomopolymers . However, short oligonucleotides {(dT)12, (dA)14} stimulate neither the protease nor the ATPase activities of the recA+ protein . In contrast, these short oligonucleotides activate tif(recA) protein to cleave lambda repressor without stimulating its ATPase activity . Moreover, both the ATPase and protease activities of the tif(recA) protein are stimulated by poly(rU) and poly(rC) whereas the recA+ protein does not respond to these ribopolymers . We have purified the recA protein from a strain in which the tif mutation is intragenically suppressed . This mutant protein (recA629) is inactive in the presence of (dT)12, (dA)14, poly(rU), and poly(rC) for lambda repressor cleavage and ATP hydrolysis . These results argue that the tif-1 mutation (or mutations) alters the DNA binding site of the recA protein . We suggest that in vivo the tif(recA) protein is activated for cleaving repressors of SOS genes by complex formation with short single-stranded regions or gaps that normally occur near the growing fork of replicating chromosomes and are too short for activating the recA+ enzyme . This mechanism can account for the expression of SOS functions in the absence of DNA damage in tif mutant strains. Eur J Biochem, 1981 Oct, 119(2), 381 - 7 tRNA thiolated pyrimidines as targets for near-ultraviolet-induced synthesis of guanosine tetraphosphate in Escherichia coli; Thomas G et al.; Illumination with near-ultraviolet light triggers synthesis of ppGpp (guanosine 3'-diphosphate 5'-diphosphate) not only in growing Escherichia coli cells containing the putative chromophore 4-thiouridine in their tRNAs {Ramabhadran, T . V and Jagger, J . (1976) Proc . Natl Acad . Sci . USA, 73, 59--69}, but also in nuv- cells which lack 4-thiouridine . The burst of ppGpp in nuv- cells is, however, induced exclusively by light of wavelengths shorter than 350 nm . Its maximum level is half that obtained in the parental strain . This ppGpp synthesis is also under the control of the relA gene, indicating that it is due to the accumulation of uncharged tRNAs . A candidate likely to trigger this effect is a 5-methylaminomethyl-2-thiouracil residues present in the first position of the anticodon loop of tRNAGlu, tRNALys and one tRNAGln isoacceptor . In conditions in vitro, this base is highly photoreactive at wavelengths shorter than 350 nm . Furthermore, near-ultraviolet-photomodified tRNAGlu and tRNALys become poor substrates of their acylation enzyme. Eur J Biochem, 1981 Oct, 119(2), 273 - 7 Modification of tryptophanase with tetranitromethane; Nihira T et al.; Modification of apotryptophanase with tetranitromethane {C(NO2)4} resulted in a loss of enzymatic activity, whereas holotryptophanase was highly resistant against C(NO2)4-inactivation . The essential importance of the active-site-bound pyridoxal 5'-phosphate (pyridoxal-P) for the protection was confirmed by the agreement of K 1/2 (protection) (1.2 microM) for pyridoxal-P with Km (1.5 microM) in enzyme catalysis . Amino acid analyses and inactivation stoichiometry showed that modification of 1--2 tyrosyl residues per monomer caused complete inactivation . The appearance of 430-nm species upon incubation of C(NO2)4-inactivated apoenzyme with pyridoxal-P indicated that the C(NO2)4-inactivated apoenzyme could still bind the coenzyme, although an affinity of the enzyme for pyridoxal-P (Kd = 51 microM) was much lower than that of the native enzyme (Kd = 0.7 microM) . A close relationship was observed between the cofactor activity of monovalent cations and their effectiveness in the protection by pyridoxal-P: in the presence of active monovalent cations (K+, NH+4 and Rb+) pyridoxal-P could provide the protection but not in the presence of inactive cations (Li+, Na+ and Cs+) as well as in the absence of inorganic monovalent cations . From the experimental results obtained it was suggested strongly that tryptophanase has essential tyrosyl residues near the active site . The tyrosyl residues were prevented from the attack of C(NO2)4 by the active-site-bound pyridoxal-P only in the catalytically active holoenzyme. Eur J Biochem, 1981 Oct, 119(2), 245 - 9 Fragments of ribosomal protein S1 and its mutant form m1-S1 . Localization of nucleic-acid-binding domain in the middle region of S1; Subramanian AR et al.; Escherichia coli ribosomal protein S1 and its mutant, shorter, form m1-S1 were cleaved at internal methionyl residues to yield, respectively, six and five fragments of Mr ranging from 1000 to 24000 . Methods are described to isolate the fragments in pure form . Four of the fragments (designated F2a, F2b, F3 and F4) contain between 86 and 215 amino acids and are therefore as large as other ribosomal proteins . Fragment F2a, derived from the N-terminal region, has previously been shown to contain the major ribosome binding domain of S1 {S . Giorginis and A . R . Subramanian (1980) J . Mol . Biol . 141, 393--408} . Here we show that the RNA binding domain of S1 is essentially contained in F3 derived from the middle region of S1 and carrying the nonreactive--SH group . The reactive--SH group of S1, whose activity is modified by ligand binding, was localized in F2b, a fragment with little RNA binding capacity . The characteristic RNA binding domain and a weak ribosome binding domain of S1 have previously been localized in the large trypsin-resistent core S1-F1 {T . Suryanarayana and A . R . Subramanian (1979) J . Mol . Biol . 127, 41--54} . Together these data indicate that two of the key functional domain of S1 are located in two regions of the molecule separated by an open, exposed segment . The present study also revealed that the nonreactive--SH group of S1 becomes reactive in m1-S1 by the loss of the remote C-terminal region in the latter. Int J Radiat Biol Relat Stud Phys Chem Med, 1981 Oct, 40(4), 375 - 84 Conversion of potentially lethal damage to lethal damage in Escherichia coli inhibited by caffeine; Koukalova B et al.; In E . coli cells with potentially lethal damage to their DNA are starved of amino acids they rapidly lose viability (Koukalova and Kuhrova 1980) . This phenomenon is called secondary lethality (SL) . In the course of secondary lethality there is a decrease in the molecular weight of DNA to a value on average three times lower (DNA fragmentation, and part of the DNA is reduced to acid-soluble fractions (DNA degradation) . Caffeine inhibits SL, and both these processes of DNA decay, The lowest effective concentration 5 mg/ml, with maximum effect at a concentration of 10 mg/ml . SL is also inhibited by the absence of an energy source . The possible mechanism of SL is discussed on the basis of these results. J Infect Dis, 1981 Oct, 144(4), 329 - 36 Properties of reference Escherichia coli endotoxin and its phthalylated derivative in humans; Elin RJ et al.; The properties of a reference bacterial endotoxin prepared from Escherichia coli and its phthalylated derivative were studied in normal human volunteers infected intravenously with the compounds . The minimal pyrogenic dose of the reference endotoxin is about 0.1-0.5 ng/kg . The increase in white blood cell count, absolute granulocyte count, absolute immature granulocyte count, and concentrations of serum amyloid A, cortisol, and growth hormone was directly related to the concentration of reference endotoxin administered . Phthalylated reference endotoxin up to 1,000 ng/kg (at least 500 ng of the patent compound/kg) was administered to normal human volunteers without significant changes in temperature, white blood cell count, absolute granulocyte count, and concentrations of serum amyloid A, cortisol, and growth hormone . Thus, this study defines biologic properties of the new reference bacterial endotoxin in humans and demonstrates effective detoxification by phthalylation of the present compound. J Bacteriol, 1981 Oct, 148(1), 83 - 90 Role of gene fadR in Escherichia coli acetate metabolism; Maloy SR et al.; Mutants of Escherichia coli K-12 constitutive for fatty acid degradation (fadR) showed an increased rate of utilization of exogenous acetate . Acetate transport, oxidation, and incorporation into macromolecules was approximately fivefold greater in fadR mutants than fadR+ strains during growth on succinate as a carbon source . This effect was due to the elevated levels of glyoxylate shunt enzymes in fadR mutants, since (i) similar results were seen with mutants constitutive for the glyoxylate shunt enzymes (iclR), (ii) induction of the glyoxylate shunt in fadR+ strains by growth on acetate or oleate increased the rate of acetate utilization to levels comparable to those in fadR mutants, and (iii) fadR and fadR+ derivatives of mutants defective for the glyoxylate shunt enzymes showed equivalent rates of acetate utilization under these conditions . These results suggest that the operation of the glyoxylate shunt may play a significant role in the utilization of exogenous acetate by fadR mutants. J Bacteriol, 1981 Oct, 148(1), 379 - 82 The gene for ribosomal protein L31, rpmE, is located at 88.5 minutes on the Escherichia coli chromosomal linkage map; Dabbs ER; Two mutations resulting in an alteration in large-subunit ribosomal protein L31 were mapped at around 88.5 min on the Escherichia coli chromosomal linkage map . They were located between metB and argH and cotransduced over 90% with metB . These mutations were shown to define the structural gene of protein L31, rpmE. J Bacteriol, 1981 Oct, 148(1), 274 - 82 Identification of the molybdenum cofactor in chlorate-resistant mutants of Escherichia coli; Amy NK; Experiments were performed to determine whether defects in molybdenum cofactor metabolism were responsible for the pleiotropic loss of the molybdoenzymes nitrate reductase and formate dehydrogenase in chl mutants of Escherichia coli . In wild-type E . coli, molybdenum cofactor activity was present in both the soluble and membrane-associated fractions when the cells were grown either aerobically or anaerobically, with and without nitrate . Molybdenum cofactor in the soluble fraction decreased when the membrane-bound nitrate reductase and formate dehydrogenase were induced . In the chl mutants, molybdenum cofactor activity was found in the soluble fraction of chlA, chlB, chlC, chlD, chlE, and chlG, but only chlB, chlC, chlD, and chlG expressed cofactor activity in the membrane fraction . The defect in the chlA mutants which prevented incorporation of the soluble cofactor into the membrane also caused the soluble cofactor to be defective in its ability to bind molybdenum . This cofactor was not active in the absence of molybdate, and it required at least threefold more molybdate than did the wild type in the Neurospora crassa nit-1 complementation assay . However, the cofactor from the chlA strain mediated the dimerization of the nit-1 subunits in the presence and absence of molybdate to yield the 7.9S dimer . Growth of chlA mutants in medium with increased molybdate did not repair the defect in the chlA cofactor nor restore the molybdoenzyme activities . Thus, molybdenum cofactor was synthesized in all the chl mutants, but additional processing steps may be missing in chlA and chlE mutants for proper insertion of cofactor in the membrane. J Bacteriol, 1981 Oct, 148(1), 265 - 73 Role of sulA and sulB in filamentation by lon mutants of Escherichia coli K-12; Gottesman S et al.; Cells containing the pleiotropic Escherichia coli mutation lon filament extensively and die after exposure to ultraviolet light . Outside suppressors of the ultraviolet sensitivity, called sul, have previously been described at two loci; these mutations reverse the ultraviolet sensitivity of lon strains but do not affect the mucoidal or degradation defect of these strains . An isogenic set of strains carrying combinations of lon, sulA, and sulI was constructed, and their behavior during normal growth and after ultraviolet treatment was studied . sulA mutations had no detectable phenotype in lon+ cells; the lon sulA strains filamented transiently after ultraviolet irradiation, as did lon+ sul+ cells . We found that the sulB mutation, which alters cell morphology and slows recovery from transient filamentation after ultraviolet treatment, was epistatic to both lon and sulA . Whereas sulA mutations were recessive to the wild-type allele, sulB was partially dominant . The simplest model to account for our observations is that sulA and lon participate in a pathway of filamentation independent of that which produces transient filamentation in wild-type strains; sulB product may be the target of sulA action and may play a role in normal cell division. J Bacteriol, 1981 Oct, 148(1), 1 - 9 Genetic analysis of mutants affected in the Pst inorganic phosphate transport system; Cox GB et al.; A number of mutant alleles affecting the Pst phosphate transport system have been divided into three complementation groups on the basis of constitutive alkaline phosphatase activity in appropriate partial diploid strains . The three complementation groups were represented by the alleles pstA2 and phoT32 and the newly described allele pstB401 . The two alleles phoS28 and phoS21 appeared to be polar . The phoS28 allele affected both the phoT and pstB genes but not the pstA gene, whereas the phoS21 allele appeared to be a mutation in the pstA gene exerting polar effects on both the pstB and phoT genes . It was concluded that the three genes pstA, pstB, and phoT were part of an operon and that the phosphate-binding protein was not coded for by any of these genes . The phoS gene, defined as the structural gene for the phosphate-binding protein, is also part of the operon, but the phoS28 and phoS21 alleles are not mutations in the phoS gene and were reclassified as pho-28 and pho-21 alleles . The gene order was concluded to be pstA-(pstB-phoT)-phoS, with the pstA gene promotor proximal and the direction of transcription opposite to that of the nearby unc operon. J Cell Physiol, 1981 Oct, 109(1), 161 - 9 A granulocyte-macrophage colony-stimulating factor (GM-CSF) produced by carrageenin-induced inflammatory cells of mice; Shikita M et al.; Seven days after subcutaneous injection of 2% carrageenin solution in mice, the induced inflammatory tissue was isolated and treated with 0.1% trypsin . When the dispersed cells were incubated in a nutrient medium containing 5-20% calf serum, the cells grew adhering to the culture-dish surface and colony-stimulating factor (CSF) was accumulated in the medium gradually . Addition of lipopolysaccharide (Escherichia coli endotoxin) in the cell culture did not enhance the CSF production . It was shown by isoelectrofocusing that the majority of the produced CSF was an acidic molecule (pI = 3.8), while the treatment of this CSF with neuraminidase yielded a less acidic CSF species (pI = 5.1) . Upon gel-filtration chromatography in the presence of 6 M guanidine HCl, the CSF exhibited an apparent molecular weight of 42,000 . On the other hand, polyacrylamide gel-electrophoresis in the presence of 0.1% sodium dodecyl sulfate gave a molecular weight estimate of 65,000 . Microscopic examination of the bone marrow cell culture showed that about one-third of the colonies were granulocytic . Addition of prostaglandin E1(PGE1) in the bone marrow cell culture significantly inhibited the action of the CSF, while prostaglandin D2 was less inhibitory than PGE1 . The result suggests that the cells isolated from the inflammatory tissue are capable of producing CSF, which may have a role for proliferation and maturation of the mononuclear phagocytes at the site of inflammation. J Virol Methods, 1981 Oct, 3(3), 155 - 65 Enzyme-linked fluorescence immunoassays using beta-galactosidase and antibodies covalently bound to polystyrene plates; Neurath AR et al.; A solid-phase enzyme-linked immunoassay using a fluorogenic substrate (4-methylumbelliferyl-beta-D-galactopyranoside) was developed . Antibodies were covalently linked to glutaraldehyde-activated 96-well aminopolystyrene plates . Antigens from test samples were adsorbed to the solid phase and detected using antibodies conjugated with E . coli beta-galactosidase . Glutaraldehyde, N-succinimidyl-3-(2-pyridyldithio)-propionate or N-succinimidyl-6-(4-azido-2-nitrophenylamino)-hexanoate were used as linkers between antibodies and the enzyme . The measurement of fluorescence can be automated for rapid screening of many specimens . The sensitivity limit of the test for HBsAg is about 5-10 pg. Gene, 1981 Oct, 15(1), 67 - 80 Cloning and characterization of nine heat-shock-induced mRNAs of Drosophila melanogaster; Lis JT et al.; We have cloned cDNA segments representing nine different genes whose expression is activated by heat-shock treatment of Drosophila melanogaster cells . These nine were selected from a larger population based on the fact that the levels of the polysomal, poly(A)-containing RNA homologous to each of these cDNA segments is significantly greater in induced cells . Among these genes are four that hybridize in situ to major heat-shock puff sites on polytene chromosomes, and five previously uncharacterized genes that hybridize to other sites . Some of these additional sites have been previously implicated as responders to heat shock . Hybridization of in vivo pulse-labeled . total RNA from induced and uninduced cells to specific cDNA segments provided an estimate of both the relative level of different heat-shock transcripts and the inducibility of the nine genes . Although the most abundant RNA species are derived from genes at major heat-shock puff sites, all nine show significant induction . These results indicate that the repertoire of gene expression that is activated or amplified by heat shock is not confined to these major puff sites and that the number of genes involved in the heat-shock response is much larger than the seven commonly noted. Pharmacol Biochem Behav, 1981 Oct, 15(4), 657 - 8 Body temperature of rats after prostaglandins E2 and F2 alpha and their precursor; Splawinski J et al.; Prostaglandins E2 and F2 alpha and arachidonic acid were injected into various brain structures outside the hypothalamus in rats . The body temperature rise induced by PGE2, but not by PGF2 alpha or arachidonic acid, was probably due to its diffusion or transport into the hypothalamus . Comparison of the latencies of the hyperthermic response to PGE2, PGF2 alpha, and arachidonic acid revealed that PGE2 was the most rapidly acting agent in structures located close to the hypothalamus . In contrast, arachidonic acid acted most rapidly after administration into the nucleus reticularis pontis caudalis. J Bacteriol, 1981 Oct, 148(1), 203 - 9 Exogenous induction of the Escherichia coli hexose phosphate transport system defined by uhp-lac operon fusions; Shattuck-Eidens DM et al.; The uhp-coded hexose phosphate transport system of Escherichia coli is normally induced by the presence of extracellular glucose-6-phosphate (G6P), whereas internally generated G6P does not provide a regulatory signal . Strains carrying uhp-lac operon fusions in which lac operon expression is under the control of the uhpT promoter were isolated . The direction of transcription of the uhp T gene was found to be counterclockwise on the E . coli chromosome map . The effects of added sugar phosphates on induction of beta-galactosidase and G6P uptake activities were compared in two fusion-carrying strains differing only in the presence of functional Uhp+ activity . Induction of uhp expression by G6P was equally effective in the two strains; accumulation of G6P diminished its ability to serve as an inducer . Mannose-6-phosphate was an effective competitive inhibitor of G6P uptake, but did not inhibit induction by G6P of uhp expression . No sugar phosphates were found that inhibited induction by G6P . Inorganic phosphate competitively inhibited induction by G6P whether G6P transport activity was present or not . Thus, the transport activity is not involved in the regulation of its synthesis, and these results strongly suppor