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In vivo cloning of a carboxy-terminalrpoB allele which confers altered transcriptional properties

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Abstract

A 3′-terminal mutation of the gene encoding the β subunit ofEscherichia coli RNA polymerase was isolated using anin vivo polA(Ts) technique. Cloning of the allele was monitored by virtue of the fact that the deletion Δ(rpoB) 1570-1 resulted in an altered-size restriction fragment. DNA sequencing confirmed the predicted nature and location of the mutation: Δ(rpoB) 1570-1 involved an in-frame deletion of 186 bp (62 codons) encoding amino acid residues 967–1028. The phenotype conferred by Δ(rpoB) 1570-1 is discussed with respect to conserved domains within the β polypeptide.

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References

  • Bedwell D.M., Nomura M.: Feedback regulation of RNA polymerase subunit synthesis after the conditional overproduction of RNA polymerase inEscherichia coli.Mol. Gen. Genet. 204, 17–23 (1986).

    Article  PubMed  CAS  Google Scholar 

  • Birnboim H.C., Doly J.: A rapid alkaline extraction procedure for screening recombinant plasmid DNA.Nucl. Acid Res. 7, 1513–1523 (1979).

    Article  CAS  Google Scholar 

  • Borukhov S., Severinov K., Kashlev M., Lebedev A., Bass I., Rowland G.C., Lim P.-P., Glass R.E., Nikiforov V., Goldfarb A.: Mapping of trypsin cleavage and antibody-binding sites and delineation of a dispensable domain in the β subunit ofEscherichia coli RNA polymerase.J. Biol. Chem. 266, 23921–23926 (1991).

    PubMed  CAS  Google Scholar 

  • Brosius J., Cate R.L., Perlmutter A.P.: Precise location of two promoters for the β lactamase gene of pBR322.J. Biol. Chem. 257, 9205–9210 (1982).

    PubMed  CAS  Google Scholar 

  • Corpet F.: Multiple sequence alignment with hierarchical clustering.Nucl. Acid Res. 16, 10881–10890 (1988).

    Article  CAS  Google Scholar 

  • Feinberg A.P., Vogelstein B.: A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity.Analyt. Biochem. 132, 6–13 (1983).

    Article  PubMed  CAS  Google Scholar 

  • Fiil N.P., Bendiak D., Collins J., Freisen J.D.: Expression ofEscherichia coli ribosomal protein and RNA polymerase genes cloned on plasmids.Mol. Gen. Genet. 173, 39–50 (1979).

    Article  PubMed  CAS  Google Scholar 

  • Glass R.E., Honda A., Ishihama A.: Genetic studies on the β subunit ofEscherichia coli RNA polymerase. IX. The role of the carboxy-terminus in enzyme assembly.Mol. Gen. Genet. 203, 492–495 (1986a).

    Article  PubMed  CAS  Google Scholar 

  • Glass R.E., Jones S.T., Ishihama A.: Genetic studies on the β subunit ofEscherichia coli RNA polymerase. VII. RNA polymerase is a target for ppGpp.Mol. Gen. Genet. 203, 265–268 (1986b).

    Article  PubMed  CAS  Google Scholar 

  • Glass R.E., Jones S.T., Ishihama A.: RNA polymerase is the target for ppGpp, pp. 393–395 inRNA Polymerase and the Regulation of Transcription (W. Reznikoff, R. Burgess, J. Dahlberg, C. Gross, M. Record, M. Wickens, Eds). Elsevier, New York 1987.

    Google Scholar 

  • Glass R.E., Ralphs N.T., Fujita N., Ishihama A.: Assembly of amber fragments of the β subunit ofEscherichia coli RNA polymerase.Eur. J. Biochem. 176, 403–407 (1988).

    Article  PubMed  CAS  Google Scholar 

  • Glickman B.W., Ripley L.S.: Structural intermediates of deletion mutagenesis: A role for palindromic DNA.Proc. Nat. Acad. Sci. 81, 512–516 (1984).

    Article  PubMed  CAS  Google Scholar 

  • Gutterson N.I., Koshland D.E.: Replacement and amplification of bacterial genes with sequences alteredin vitro.Proc. Nat. Acad. Sci. 80, 4894–4898 (1983).

    Article  PubMed  CAS  Google Scholar 

  • Ishihama A., Fujita N., Glass R.E.: Subunit assembly and metabolic stability ofE. coli RNA polymeraseProteins: Structure Function and Genetics 2, 42–53 (1987).

    Article  CAS  Google Scholar 

  • Jin D.J., Gross C.A.: ThreerpoBC mutations that suppress the termination defects ofrho mutants also affect the functions ofnusA mutants.Mol. Gen. Genet. 216, 269–275 (1989).

    Article  PubMed  CAS  Google Scholar 

  • Jin D.J., Cashel M., Friedman D.I., Nakamura Y., Walter W.A., Gross C.A.: Effects of rifampicin resistantrpoB mutations on antitermination and interaction withnusA inEscherichia coli.J. Mol. Biol. 204, 247–261 (1988a).

    Article  PubMed  CAS  Google Scholar 

  • Jin D.J., Walter W.A., Gross C.A.: Characterisation of the termination phenotypes of rifampicin-resistant mutants.J. Mol. Biol. 202, 245–253 (1988b).

    Article  PubMed  CAS  Google Scholar 

  • Jones I.M., Primrise S.B., Ehrlich S.D.: Recombination between short direct repeats in therecA host.Mol. Gen. Genet. 188, 486–489 (1982).

    Article  PubMed  CAS  Google Scholar 

  • Listtisyn N.A., Monastyrskaya G.S., Sverdlov E.D.: Genes coding for RNA polymerase β subunit in bacteriaEur. J. Biochem. 177, 363–369 (1989).

    Article  Google Scholar 

  • Mead D.A., Szczesna-Skorupa E., Kemper B.: Single-stranded DNA ‘blue’ T7 promoter plasmids: a versatile tandem promoter system for cloning and protein engineering.Prot. Eng. 1, 67–74 (1986).

    Article  CAS  Google Scholar 

  • Meek D.W., Hayward R.S.: Direct evidence for autogenous regulation of theEscherichia coli genesrpoBC in vivo Mol. Gen. Genet. 202, 500–508 (1986).

    Article  PubMed  CAS  Google Scholar 

  • Nene V., Glass R.E.: Non-essential sequences in the β subunit ofE. coli RNA polymerase pp. 173–181 inGene Manipulation and Expression (R.E. Glass, J. Spižek, Eds). Croom Helm 1984a.

  • Nene V., Glass R.E.: Genetic studies on the β subunit ofEscherichia coli RNA polymerase. IV. Structure— function correlates.Mol. Gen. Genet. 194, 166–172 (1984b).

    Article  PubMed  CAS  Google Scholar 

  • Nene V., Glass R.E.: Genetic studies on the β subunit ofEscherichia coli RNA polymerase. VI. A redundant region in the β polypeptide.Mol. Gen. Genet. 196, 64–67 (1984c).

    Article  PubMed  CAS  Google Scholar 

  • Nikiforov V.G., Kalyaeva E.S., Moiseyeva E.P., Yakubov L.Z.: Functionally important site in the vicinity of the aminoterminus of theE. coli RNA polymerase β subunit.FEBS Lett. 191, 72–74 (1985).

    Article  PubMed  CAS  Google Scholar 

  • Rowland G.C., Glass R.E.: Conservation of RNA polymerase.Bioessays 12, 343–346 (1990).

    Article  PubMed  CAS  Google Scholar 

  • Sambrook J., Fritsch E.F., Maniatis T.:Molecular Cloning. A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor (New York) 1989.

    Google Scholar 

  • Sanger F., Nicklen S., Coulson A.R.: DNA sequencing with chain-terminating inhibitors.Proc. Nat. Acad. Sci. 74, 5463–5467 (1977).

    Article  PubMed  CAS  Google Scholar 

  • Silhavy T.J., Berman M.L., Enquist L.W.:Experiments with Gene Fusions. Cold Spring Harbor (New York) 1984.

  • Southern E.M.: Detection of specific sequences among DNA fragments separated by gel electrophoresis.J. Mol. Biol. 98, 503–517 (1975).

    Article  PubMed  CAS  Google Scholar 

  • Tittawella, I.P.B.: Evidence for clustering of RNA polymerase and ribosomal protein genes in species of Enterobacteria.Mol. Gen. Genet. 195, 215–218 (1984).

    Article  PubMed  CAS  Google Scholar 

  • Woese C.R.: Bacterial Evolution.Microbiol. Rev. 51, 221–271 (1987).

    PubMed  CAS  Google Scholar 

  • Yura T., Ishihama A.: Genetics of bacterial RNA polymerases.Ann. Rev. Genet. 13, 59–97 (1979)

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Biochemistry, University of Nottingham Medical School, Queen's Medical Centre, NG7 2UH, Nottingham, UK

    G. C. Rowland, P. -P. Lim & R. E. Glass

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  1. G. C. Rowland
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  2. P. -P. Lim
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  3. R. E. Glass
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Dedicated to Dr. J. Spížek on the occasion of his 60th birthday

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Rowland, G.C., Lim, P.P. & Glass, R.E. In vivo cloning of a carboxy-terminalrpoB allele which confers altered transcriptional properties. Folia Microbiol 40, 588–594 (1995). https://doi.org/10.1007/BF02818514

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