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. 1982 Aug;151(2):609–619. doi: 10.1128/jb.151.2.609-619.1982

Mutations affecting regulation of methionine biosynthetic genes isolated by use of met-lac fusions.

J T Mulligan, W Margolin, J H Krueger, G C Walker
PMCID: PMC220301  PMID: 6284705

Abstract

Fusions of the lac genes to the promoters of four structural genes in the methionine biosynthetic pathway, metA, metB, metE, and metF, were obtained by the use of the Mu d(Ap lac) bacteriophage. The levels of beta-galactosidase in these strains could be derepressed by growth under methionine-limiting conditions. Furthermore, growth in the presence of vitamin B12 repressed the synthesis of beta-galactosidase in strains containing a fusion of lacZ to the metE promoter, phi(metE'-lacZ+). Mutations affecting the regulation of met-lac fusions were generated by the insertion of Tn5. Tn5 insertions were obtained at the known regulatory loci metJ and metK. Interestingly, a significant amount of methionine adenosyltransferase activity remained in the metK mutant despite the fact that the mutation was generated by an insertion. Several Tn5-induced regulatory mutations were isolated by screening for high-level beta-galactosidase expression in a phi(metE'-lacZ+) strain in the presence of vitamin B12. Tn5 insertions mapping at the btuB (B12 uptake), metH (B12 dependent tetrahydropteroylglutamate methyltransferase), and metF (5,10-methylenetetrahydrofolate reductase) loci were obtained. The isolation of the metH mutant was consistent with previous suggestions that the metH gene product is required for the repression of metE by vitamin B12. The metF::Tn5 insertion was of particular interest since it suggested that a functional metf gene product was also needed for repression of metE by vitamin B12.

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Selected References

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

  1. Ahmed A. Mechanism of repression of methionine biosynthesis in Escherichia coli. I. The role of methionine, s-adenosylmethionine, and methionyl-transfer ribonucleic acid in repression. Mol Gen Genet. 1973 Jul 16;123(4):299–324. doi: 10.1007/BF00433648. [DOI] [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. Bassford P. J., Jr, kadner R. J. Genetic analysis of components involved in vitamin B12 uptake in Escherichia coli. J Bacteriol. 1977 Dec;132(3):796–805. doi: 10.1128/jb.132.3.796-805.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Campbell A., Berg D. E., Botstein D., Lederberg E. M., Novick R. P., Starlinger P., Szybalski W. Nomenclature of transposable elements in prokaryotes. Gene. 1979 Mar;5(3):197–206. doi: 10.1016/0378-1119(79)90078-7. [DOI] [PubMed] [Google Scholar]
  5. Casadaban M. J., Cohen S. N. Lactose genes fused to exogenous promoters in one step using a Mu-lac bacteriophage: in vivo probe for transcriptional control sequences. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4530–4533. doi: 10.1073/pnas.76.9.4530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Casadaban M. J. Transposition and fusion of the lac genes to selected promoters in Escherichia coli using bacteriophage lambda and Mu. J Mol Biol. 1976 Jul 5;104(3):541–555. doi: 10.1016/0022-2836(76)90119-4. [DOI] [PubMed] [Google Scholar]
  7. Cherest H., Surdin-Kerjan Y. S-adenosyl methionine requiring mutants in Saccharomyces cerevisiae: evidences for the existence of two methionine adenosyl transferases. Mol Gen Genet. 1978 Jul 11;163(2):153–167. doi: 10.1007/BF00267406. [DOI] [PubMed] [Google Scholar]
  8. Dawes J., Foster M. A. Vitamin B 12 and methionine synthesis in Escherichia coli. Biochim Biophys Acta. 1971 Jun 22;237(3):455–464. doi: 10.1016/0304-4165(71)90263-7. [DOI] [PubMed] [Google Scholar]
  9. Di Masi D. R., White J. C., Schnaitman C. A., Bradbeer C. Transport of vitamin B12 in Escherichia coli: common receptor sites for vitamin B12 and the E colicins on the outer membrane of the cell envelope. J Bacteriol. 1973 Aug;115(2):506–513. doi: 10.1128/jb.115.2.506-513.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Greene R. C., Hunter J. S., Coch E. H. Properties of metK mutants of Escherichia coli K-12. J Bacteriol. 1973 Jul;115(1):57–67. doi: 10.1128/jb.115.1.57-67.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Greene R. C. Methionine limitation in Escherichia coli K-12 by growth on the sulfoxides of D-methionine. J Bacteriol. 1973 Oct;116(1):230–234. doi: 10.1128/jb.116.1.230-234.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Greene R. C., Su C. H., Holloway C. T. S-Adenosylmethionine synthetase deficient mutants of Escherichia coli K-12 with impaired control of methionine biosynthesis. Biochem Biophys Res Commun. 1970 Mar 27;38(6):1120–1126. doi: 10.1016/0006-291x(70)90355-4. [DOI] [PubMed] [Google Scholar]
  13. Greene R. C., Williams R. D., Kung H. F., Spears C., Weissbach H. Effect of methionine and vitamin B-12 on the activities of methionine biosynthetic enzymes in metJ mutants of Escherichia coli K12. Arch Biochem Biophys. 1973 Sep;158(1):249–256. doi: 10.1016/0003-9861(73)90619-x. [DOI] [PubMed] [Google Scholar]
  14. Hafner E. W., Tabor C. W., Tabor H. Isolation of a metK mutant with a temperature-sensitive S-adenosylmethionine synthetase. J Bacteriol. 1977 Dec;132(3):832–840. doi: 10.1128/jb.132.3.832-840.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Herman G. E., Modrich P. Escherichia coli K-12 clones that overproduce dam methylase are hypermutable. J Bacteriol. 1981 Jan;145(1):644–646. doi: 10.1128/jb.145.1.644-646.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hobson A. C., Smith D. A. S-adenosylmethionine synthetase in methionine regulatory mutants of Salmonella typhimurium. Mol Gen Genet. 1973 Oct 16;126(1):7–18. doi: 10.1007/BF00333477. [DOI] [PubMed] [Google Scholar]
  17. Holloway C. T., Greene R. C., Su C. H. Regulation of S-adenosylmethionine synthetase in Escherichia coli. J Bacteriol. 1970 Nov;104(2):734–747. doi: 10.1128/jb.104.2.734-747.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hunter J. S., Greene R. C., Su C. H. Genetic characterization of the metK locus in Escherichia coli K-12. J Bacteriol. 1975 Jun;122(3):1144–1152. doi: 10.1128/jb.122.3.1144-1152.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Komeda Y., Iino T. Regulation of expression of the flagellin gene (hag) in Escherichia coli K-12: analysis of hag-lac gene fusions. J Bacteriol. 1979 Sep;139(3):721–729. doi: 10.1128/jb.139.3.721-729.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Krueger J. H., Johnson J. R., Greene R. C., Dresser M. Structural studies of lambda transducing bacteriophage carrying bacterial deoxyribonucleic acid from the metBJLF region of the Escherichia coli chromosome. J Bacteriol. 1981 Aug;147(2):612–621. doi: 10.1128/jb.147.2.612-621.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kung H. F., Spears C., Greene R. C., Weissbach H. Regulation of the terminal reactions in methionine biosynthesis by vitamin B 12 and methionine. Arch Biochem Biophys. 1972 May;150(1):23–31. doi: 10.1016/0003-9861(72)90005-7. [DOI] [PubMed] [Google Scholar]
  22. Marinus M. G., Morris N. R. Isolation of deoxyribonucleic acid methylase mutants of Escherichia coli K-12. J Bacteriol. 1973 Jun;114(3):1143–1150. doi: 10.1128/jb.114.3.1143-1150.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Milner L., Whitfield C., Weissbach H. Effect of L-methionine and vitamin B 12 on methionine biosynthesis in Escherichia coli. Arch Biochem Biophys. 1969 Sep;133(2):413–419. doi: 10.1016/0003-9861(69)90470-6. [DOI] [PubMed] [Google Scholar]
  24. Su C. H., Greene R. C. Regulation of methionine biosynthesis in Escherichia coli: mapping of the metJ locus and properties of a metJ plus-metJ minus diploid. Proc Natl Acad Sci U S A. 1971 Feb;68(2):367–371. doi: 10.1073/pnas.68.2.367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Söll D. Enzymatic modification of transfer RNA. Science. 1971 Jul 23;173(3994):293–299. doi: 10.1126/science.173.3994.293. [DOI] [PubMed] [Google Scholar]
  26. TABOR H., ROSENTHAL S. M., TABOR C. W. The biosynthesis of spermidine and spermine from putrescine and methionine. J Biol Chem. 1958 Oct;233(4):907–914. [PubMed] [Google Scholar]
  27. Yamamoto M., Nomura M. Isolation of lambda transducing phages carrying rRNA genes at the metA-purD region of the Escherichia coli chromosome. FEBS Lett. 1976 Dec 31;72(2):256–261. doi: 10.1016/0014-5793(76)80981-7. [DOI] [PubMed] [Google Scholar]

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