Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1982 Aug;151(2):899–904. doi: 10.1128/jb.151.2.899-904.1982

Isolation of single-site Escherichia coli mutants deficient in thiamine and 4-thiouridine syntheses: identification of a nuvC mutant.

J Ryals, R Y Hsu, M N Lipsett, H Bremer
PMCID: PMC220340  PMID: 6178725

Abstract

A method is described to rapidly select and classify many independent near-UV irradiation-resistant Escherichia coli mutants, which include tRNA modification and RNA synthesis control mutants. One class of these mutants was found to be simultaneously deficient in thiamine biosynthesis and in the ability to modify uridine in tRNA to 4-thiouridine, known to be the target for near-UV irradiation. These mutants were found to be unable to make thiazole, a thiamine precursor. The addition of thiazole restores the thiamine deficiency but does not render the cells near-UV irradiation sensitive. In vitro studies on one of these mutants indicated a deficiency in protein factor C (nuvC), required for the 4-thiouridine modification of tRNA. In P1 transduction, the thiazole marker cotransduced with the histidine marker, which places the thiazole marker between 42 and 46 min on the E. coli chromosome map. Both thiamine production and 4-thiouridine production were resumed by 87% of the spontaneous reversions, suggesting a single-point mutation. Our results indicate that we have isolated nuvC mutants and that the nuvC polypeptide is involved in two functions, tRNA modification and thiazole biosynthesis.

Full text

PDF
901

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Abrell J. W., Kaufman E. E., Lipsett M. N. The biosynthesis of 4-thiouridylate. Separation and purification of two enzymes in the transfer ribonucleic acid-sulfurtransferase system. J Biol Chem. 1971 Jan 25;246(2):294–301. [PubMed] [Google Scholar]
  2. Bachmann B. J., Low K. B. Linkage map of Escherichia coli K-12, edition 6. Microbiol Rev. 1980 Mar;44(1):1–56. doi: 10.1128/mr.44.1.1-56.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Jagger J. Growth delay and photoprotection induced by near-ultraviolet light. Res Prog Org Biol Med Chem. 1972;3(Pt 1):383–401. [PubMed] [Google Scholar]
  4. Kubitschek H. E. Mutagenesis by near-visible light. Science. 1967 Mar 24;155(3769):1545–1546. doi: 10.1126/science.155.3769.1545. [DOI] [PubMed] [Google Scholar]
  5. Lipsett M. N. Enzymes producing 4-thiouridine in Escherichia coli tRNA: approximate chromosomal locations of the genes and enzyme activities in a 4-thiouridine-deficient mutant. J Bacteriol. 1978 Sep;135(3):993–997. doi: 10.1128/jb.135.3.993-997.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Peterkofsky A., Lipsett M. N. The origin of the sulfur in s-RNA. Biochem Biophys Res Commun. 1965 Sep 22;20(6):780–786. doi: 10.1016/0006-291x(65)90086-0. [DOI] [PubMed] [Google Scholar]
  7. Ramabhadran T. V., Fossum T., Jagger J. Escherichia coli mutant lacking 4-thiouridine in its transfer ribonucleic acid. J Bacteriol. 1976 Nov;128(2):671–672. doi: 10.1128/jb.128.2.671-672.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ramabhadran T. V., Fossum T., Jagger J. In vivo induction of 4-thiouridine-cytidine adducts in tRNA of E. coli B/r by near-ultraviolet radiation. Photochem Photobiol. 1976 May;23(5):315–321. doi: 10.1111/j.1751-1097.1976.tb07254.x. [DOI] [PubMed] [Google Scholar]
  9. Ramabhadran T. V., Jagger J. Mechanism of growth delay induced in Escherichia coli by near ultraviolet radiation. Proc Natl Acad Sci U S A. 1976 Jan;73(1):59–63. doi: 10.1073/pnas.73.1.59. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Ramabhadran T. V. Method for the isolation of Escherichia coli relaxed mutants, utilizing near-ultraviolet irradiation. J Bacteriol. 1976 Sep;127(3):1587–1589. doi: 10.1128/jb.127.3.1587-1589.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. STENT G. S., BRENNER S. A genetic locus for the regulation of ribonucleic acid synthesis. Proc Natl Acad Sci U S A. 1961 Dec 15;47:2005–2014. doi: 10.1073/pnas.47.12.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Yaniv M., Favre A., Barrell B. G. Structure of transfer RNA. Evidence for interaction between two non-adjacent nucleotide residues in tRNA from Escherichia coli. Nature. 1969 Sep 27;223(5213):1331–1333. doi: 10.1038/2231331a0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES