Molecular and General Genetics MGG

, Volume 209, Issue 3, pp 481–488

Cloning and nucleotide sequencing of the genes rimI and rimJ which encode enzymes acetylating ribosomal proteins S18 and S5 of Escherichia coli K12

  • Akikazu Yoshikawa
  • Setsuko Isono
  • Abraham Sheback
  • Katsumi Isono
Article

Summary

The rimI gene of Escherichia coli K12, which encodes an enzyme catalysing acetylation of the N-terminal alanine of ribosomal protein S18, has been cloned into a mini-F plasmid pRE432 and characterized at the molecular level. Similarly, the rimJ gene, which encodes another acetylating enzyme that is specific for ribosomal protein S5, has been cloned and characterized. From the nucleotide sequence data for the two genes the RimI enzyme was deduced to contain 161 amino acid residues with a calculated molecular weight (Mr) of 18232 and the RimJ enzyme contains 194 amino acid residues with a calculated Mr of 22687. The proteins produced from the two genes in maxi-cells were identified by electrophoresis on acrylamide gels and their operon structure was analysed by insertional mutagenesis with transposon γδ (Tn1000) and by measuring the size of their transcripts. Their structural homology was analysed by DNA hybridization and by calculation with computer programs. There is only a low level of overall homology between the two genes except for a 3′ terminal region in which a significant degree of homology was noticed.

Key words

N-terminal acetylation Ribosomal proteins Molecular cloning Nucleotide sequencing 

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References

  1. Bochner BR, Huang H-C, Schieven GL, Ames BN (1980) Positive selection for loss of tetracyclin resistance. J Bacteriol 143:926–933Google Scholar
  2. Colson C, Lhoest J, Urlings C (1979) Genetics of ribosomal protein methylation in Escherichia coli. III. Map position of two genes, prmA and prmB, governing methylation of protein L11 and L3. Mol Gen Genet 169:245–250Google Scholar
  3. Cumberlidge AG, Isono K (1979) Ribosomal protein modification in Escherichia coli. I. A mutant lacking the N-terminal acetylation of protein S5 exhibits thermosensitivity. J Mol Biol 131:169–189Google Scholar
  4. Favaloro J, Treisman R, Kamen R (1980) Transcription maps of polyoma virus-specific RNA: Analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol 65:718–749Google Scholar
  5. Geyl D, Böck A, Isono K (1981) An improved method for two-dimensional gel-electrophoresis: Analysis of mutationally altered ribosomal proteins of Escherichia coli. Mol Gen Genet 181:309–312Google Scholar
  6. Guyer MS (1983) Uses of the transposon γδ in the analysis of cloned genes. Methods Enzymol 101:362–369Google Scholar
  7. Hawley DK, McClure WR (1983) Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res 11:2237–2255Google Scholar
  8. Imamoto F (1969) Intragenic initiations of transcription of the tryptophan operon in Escherichia coli following dinitrophenol treatment without tryptophan. J Mol Biol 43:51–69Google Scholar
  9. Isono K (1984) A computer program package for storing and retrieving DNA/RNA and protein sequence data. Nucleic Acids Res 12:101–112Google Scholar
  10. Isono K, Isono S (1980) Ribosomal protein modification in Escherichia coli. II Studies of a mutant lacking the N-terminal acetylation of protein S18. Mol Gen Genet 177:645–651Google Scholar
  11. Janda I, Kitakawa M, Isono K (1985) Gene rpmF for ribosomal protein L32 and gene rimJ for a ribosomal protein acetylating enzyme are located near pyrC (23.4 min) in Escherichia coli. Mol Gen Genet 201:433–436Google Scholar
  12. Lindahl L, Zengel JM (1986) Ribosomal genes in Escherichia coli. Annu Rev Genet 20:297–326Google Scholar
  13. Maloy SR, Nunn WD (1981) Selection for loss of tetracycline resistance by Escherichia coli. J Bacteriol 145:1110–1112Google Scholar
  14. Maniatis T, Fritsch EF, Sambrook J (1982) Molecular Cloning: A laboratory manual. Cold Spring Harbour Laboratory, Cold Spring Harbor, New YorkGoogle Scholar
  15. Messing J (1983) New M13 vector for cloning. Methods Enzymol 101:20–78Google Scholar
  16. Mizusawa S, Nishimura S, Seela F (1986) Improvement of the dideoxy chain termination method of DNA sequencing by use of deoxy-7-deazaguanosine triphosphate in place of dGTP. Nucleic Acids Res 14:1319–1324Google Scholar
  17. O'Farrell PH, Kutter E, Nakanishi M (1980) A restriction map of the bacteriophage T4 genome. Mol Gen Genet 179:421–435Google Scholar
  18. Reeh S, Pedersen S (1979) Post-translational modification of Escherichia coli ribosomal protein S6. Mol Gen Genet 173:183–187Google Scholar
  19. Sancar A, Hack AM, Rupp WD (1979) Simple method for identification of plasmid-coded proteins. J Bacteriol 137:692–693Google Scholar
  20. Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chainterminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467Google Scholar
  21. Southern EM (1975) Detection of specific sequences among the DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517Google Scholar
  22. Wittmann HG, Littlechild JA, Wittmann-Liebold B (1979) Structure of ribosomal proteins. In: Chambliss G, Craven GR, Davies J, Davis K, Kahan L, Nomura M (eds) Ribosomes: Structure, function and genetics. University Park Press, Baltimore, pp 51–88Google Scholar
  23. Yanisch-Perron C, Vieira J, Messing J (1985) Improved M13 phage cloning vectors and host strains: Nucleotide sequences of the M13 mp18 and pUC19 vectors. Gene 33:103–119Google Scholar

Copyright information

© Springer-Verlag 1987

Authors and Affiliations

  • Akikazu Yoshikawa
    • 1
  • Setsuko Isono
    • 1
  • Abraham Sheback
    • 2
  • Katsumi Isono
    • 1
  1. 1.Department of Biology, Faculty of ScienceKobe UniversityKobeJapan
  2. 2.Abteilung WittmannMax-Planck-Institut für Molekulare GenetikBerlin 33

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