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. 1997 Oct 15;25(20):4093–4097. doi: 10.1093/nar/25.20.4093

The spoU gene of Escherichia coli, the fourth gene of the spoT operon, is essential for tRNA (Gm18) 2'-O-methyltransferase activity.

B C Persson 1, G Jäger 1, C Gustafsson 1
PMCID: PMC146995  PMID: 9321663

Abstract

We have evidence that the open reading frame previously denoted spoU is necessary for tRNA (Gm18) 2'-O-methyltransferase activity. The spoU gene is located in the gmk-rpoZ-spoT-spoU-recG operon at 82 minutes on the Escherichia coli chromosome. The deduced amino acid sequence of spoU shows strong similarities to previously characterized 2'-O-methyltransferases. Comparison of the nucleoside modification pattern of hydrolyzed tRNA, 16S rRNA and 23S rRNA from wild-type and spoU null mutants showed that the modified nucleoside 2'-O-methylguanosine (Gm), present in a subset of E. coli tRNAs at residue 18, is completely absent in the spoU mutant, suggesting that spoU encodes tRNA (Gm18) 2'-O-methyltransferase. Nucleoside modification of 16S and 23S rRNA was unaffected in the spoU mutant. Insertions in the downstream recG gene did not affect RNA modification. Absence of Gm18 in tRNA does not influence growth rate under the tested conditions and does not interfere with activity of the SupF amber suppressor, a suppressor tRNA that normally has the Gm18 modification. We suggest that the spoU gene be renamed trmH (tRNA methylation).

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Selected References

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  1. Agris P. F., Koh H., Söll D. The effect of growth temperatures on the in vivo ribose methylation of Bacillus stearothermophilus transfer RNA. Arch Biochem Biophys. 1973 Jan;154(1):277–282. doi: 10.1016/0003-9861(73)90058-1. [DOI] [PubMed] [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  3. Aström S. U., Byström A. S. Rit1, a tRNA backbone-modifying enzyme that mediates initiator and elongator tRNA discrimination. Cell. 1994 Nov 4;79(3):535–546. doi: 10.1016/0092-8674(94)90262-3. [DOI] [PubMed] [Google Scholar]
  4. BERTANI G. Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli. J Bacteriol. 1951 Sep;62(3):293–300. doi: 10.1128/jb.62.3.293-300.1951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bachmann B. J. Linkage map of Escherichia coli K-12, edition 8. Microbiol Rev. 1990 Jun;54(2):130–197. doi: 10.1128/mr.54.2.130-197.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bibb M. J., Bibb M. J., Ward J. M., Cohen S. N. Nucleotide sequences encoding and promoting expression of three antibiotic resistance genes indigenous to Streptomyces. Mol Gen Genet. 1985;199(1):26–36. doi: 10.1007/BF00327505. [DOI] [PubMed] [Google Scholar]
  7. Buck M., Connick M., Ames B. N. Complete analysis of tRNA-modified nucleosides by high-performance liquid chromatography: the 29 modified nucleosides of Salmonella typhimurium and Escherichia coli tRNA. Anal Biochem. 1983 Feb 15;129(1):1–13. doi: 10.1016/0003-2697(83)90044-1. [DOI] [PubMed] [Google Scholar]
  8. Gefter M. L. The in vitro synthesis of 2'-omethylguanosine and 2-methylthio 6N (gamma,gamma, dimethylallyl) adenosine in transfer RNA of Escherichia coli. Biochem Biophys Res Commun. 1969 Aug 7;36(3):435–441. doi: 10.1016/0006-291x(69)90583-x. [DOI] [PubMed] [Google Scholar]
  9. Gehrke C. W., Kuo K. C., McCune R. A., Gerhardt K. O., Agris P. F. Quantitative enzymatic hydrolysis of tRNAs: reversed-phase high-performance liquid chromatography of tRNA nucleosides. J Chromatogr. 1982 Jul 9;230(2):297–308. [PubMed] [Google Scholar]
  10. Gentry D., Bengra C., Ikehara K., Cashel M. Guanylate kinase of Escherichia coli K-12. J Biol Chem. 1993 Jul 5;268(19):14316–14321. [PubMed] [Google Scholar]
  11. Gustafsson C., Reid R., Greene P. J., Santi D. V. Identification of new RNA modifying enzymes by iterative genome search using known modifying enzymes as probes. Nucleic Acids Res. 1996 Oct 1;24(19):3756–3762. doi: 10.1093/nar/24.19.3756. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kalman M., Murphy H., Cashel M. The nucleotide sequence of recG, the distal spo operon gene in Escherichia coli K-12. Gene. 1992 Jan 2;110(1):95–99. doi: 10.1016/0378-1119(92)90449-y. [DOI] [PubMed] [Google Scholar]
  13. Kawai G., Ue H., Yasuda M., Sakamoto K., Hashizume T., McCloskey J. A., Miyazawa T., Yokoyama S. Relation between functions and conformational characteristics of modified nucleosides found in tRNAs. Nucleic Acids Symp Ser. 1991;(25):49–50. [PubMed] [Google Scholar]
  14. Kawai G., Yamamoto Y., Kamimura T., Masegi T., Sekine M., Hata T., Iimori T., Watanabe T., Miyazawa T., Yokoyama S. Conformational rigidity of specific pyrimidine residues in tRNA arises from posttranscriptional modifications that enhance steric interaction between the base and the 2'-hydroxyl group. Biochemistry. 1992 Feb 4;31(4):1040–1046. doi: 10.1021/bi00119a012. [DOI] [PubMed] [Google Scholar]
  15. Kiss-László Z., Henry Y., Bachellerie J. P., Caizergues-Ferrer M., Kiss T. Site-specific ribose methylation of preribosomal RNA: a novel function for small nucleolar RNAs. Cell. 1996 Jun 28;85(7):1077–1088. doi: 10.1016/s0092-8674(00)81308-2. [DOI] [PubMed] [Google Scholar]
  16. Koonin E. V., Rudd K. E. SpoU protein of Escherichia coli belongs to a new family of putative rRNA methylases. Nucleic Acids Res. 1993 Nov 25;21(23):5519–5519. doi: 10.1093/nar/21.23.5519. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kumagai I., Watanabe K., Oshima T. A thermostable tRNA (guanosine-2')-methyltransferase from Thermus thermophilus HB27 and the effect of ribose methylation on the conformational stability of tRNA. J Biol Chem. 1982 Jul 10;257(13):7388–7395. [PubMed] [Google Scholar]
  18. Ladner J. E., Jack A., Robertus J. D., Brown R. S., Rhodes D., Clark B. F., Klug A. Structure of yeast phenylalanine transfer RNA at 2.5 A resolution. Proc Natl Acad Sci U S A. 1975 Nov;72(11):4414–4418. doi: 10.1073/pnas.72.11.4414. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lloyd R. G., Sharples G. J. Molecular organization and nucleotide sequence of the recG locus of Escherichia coli K-12. J Bacteriol. 1991 Nov;173(21):6837–6843. doi: 10.1128/jb.173.21.6837-6843.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Miller J. H., Albertini A. M. Effects of surrounding sequence on the suppression of nonsense codons. J Mol Biol. 1983 Feb 15;164(1):59–71. doi: 10.1016/0022-2836(83)90087-6. [DOI] [PubMed] [Google Scholar]
  21. Neidhardt F. C., Bloch P. L., Smith D. F. Culture medium for enterobacteria. J Bacteriol. 1974 Sep;119(3):736–747. doi: 10.1128/jb.119.3.736-747.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Nicoloso M., Qu L. H., Michot B., Bachellerie J. P. Intron-encoded, antisense small nucleolar RNAs: the characterization of nine novel species points to their direct role as guides for the 2'-O-ribose methylation of rRNAs. J Mol Biol. 1996 Jul 12;260(2):178–195. doi: 10.1006/jmbi.1996.0391. [DOI] [PubMed] [Google Scholar]
  23. Quigley G. J., Wang A. H., Seeman N. C., Suddath F. L., Rich A., Sussman J. L., Kim S. H. Hydrogen bonding in yeast phenylalanine transfer RNA. Proc Natl Acad Sci U S A. 1975 Dec;72(12):4866–4870. doi: 10.1073/pnas.72.12.4866. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sirum-Connolly K., Mason T. L. Functional requirement of a site-specific ribose methylation in ribosomal RNA. Science. 1993 Dec 17;262(5141):1886–1889. doi: 10.1126/science.8266080. [DOI] [PubMed] [Google Scholar]
  25. Sprinzl M., Steegborn C., Hübel F., Steinberg S. Compilation of tRNA sequences and sequences of tRNA genes. Nucleic Acids Res. 1996 Jan 1;24(1):68–72. doi: 10.1093/nar/24.1.68. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Stern S., Moazed D., Noller H. F. Structural analysis of RNA using chemical and enzymatic probing monitored by primer extension. Methods Enzymol. 1988;164:481–489. doi: 10.1016/s0076-6879(88)64064-x. [DOI] [PubMed] [Google Scholar]
  27. Tycowski K. T., Smith C. M., Shu M. D., Steitz J. A. A small nucleolar RNA requirement for site-specific ribose methylation of rRNA in Xenopus. Proc Natl Acad Sci U S A. 1996 Dec 10;93(25):14480–14485. doi: 10.1073/pnas.93.25.14480. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. VOGEL H. J., BONNER D. M. Acetylornithinase of Escherichia coli: partial purification and some properties. J Biol Chem. 1956 Jan;218(1):97–106. [PubMed] [Google Scholar]
  29. Wu-Baer F., Lane W. S., Gaynor R. B. The cellular factor TRP-185 regulates RNA polymerase II binding to HIV-1 TAR RNA. EMBO J. 1995 Dec 1;14(23):5995–6009. doi: 10.1002/j.1460-2075.1995.tb00288.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Xiao H., Kalman M., Ikehara K., Zemel S., Glaser G., Cashel M. Residual guanosine 3',5'-bispyrophosphate synthetic activity of relA null mutants can be eliminated by spoT null mutations. J Biol Chem. 1991 Mar 25;266(9):5980–5990. [PubMed] [Google Scholar]

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