1887

Abstract

The genes of the biosynthetic pathway of ectoine (1,4,5,6-tetrahydro-2-methyl-4-pyrimidinecarboxylic acid) from the Gram-positive moderate halophile were cloned by functional expression in These genes were not only expressed, but also osmoregulated in as demonstrated by increasing cytoplasmic ectoine concentration in response to medium salinity. Sequencing of a 4∙4 kb fragment revealed four major ORFs, which were designated and The significance of three of these genes for ectoine synthesis was proved by sequence comparison with known proteins and by physiological experiments. Several deletion derivatives of the sequenced fragment were introduced into and the resulting clones were investigated for their ability to synthesize ectoine or one of the intermediates in its biosynthetic pathway. It was demonstrated that codes for -2,4-diaminobutyric acid acetyltransferase, for -2,4-diaminobutyric acid transaminase and for -ectoine synthase. A DNA region upstream of was shown to be necessary for the regulated expression of ectoine synthesis in response to the osmolarity of the medium.

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1997-04-01
2021-07-31
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References

  1. Altendorf K., Epstein W. Kdp-ATPase of Escherichia coli . Cell Physiol Biochem 1993; 4:160–168
    [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman E. J. Basic local alignment search tool .. J Mol Biol 1990; 215:403–410
    [Google Scholar]
  3. Ausubel F. M., Brent R., Kingston R. E., Moore D. D., Seidman J. G., Smith J. A., Struhl K. 1991; Current Protocols in Molecular Biology. New York : Wiley
    [Google Scholar]
  4. Bartsch K., von Johnn-Marteville A., Schulz A. 1990; Molecular analysis of two genes of the Escherichia col gabi cluster : nucleotide sequence of the glutamate : succinic semialdehyde transaminase gene (gabT) and characterization of the succinic semialdehyde dehydrogenase gene (gabD) .. J Bacteriol 172:7035–7042
    [Google Scholar]
  5. Bligh E. G., Dyer W. J. A rapid method of lipid extraction and purification.. Can J Biochem Physiol 1959; 37:911–917
    [Google Scholar]
  6. Booth I. R., Higgins C. F. Enteric bacteria and osmotic stress : intracellular potassium glutamate as a secondary signal of osmotic stress ?. FEMS Microbiol Rev 1990; 75:239–246
    [Google Scholar]
  7. Brown A. D. Microbial water stress. Bact Rev 1976; 40:803–846
    [Google Scholar]
  8. Cohen S. N., Chang A. C. Y., Hsu L. Non chromosomal antibiotic resistance in bacteria : genetic transformation of Escherichia coli by R-factor DNA.. Proc Natl Acad Sci USA 1972; 69:2110–2140
    [Google Scholar]
  9. Csonka L. N., Hanson A. D. Prokaryotic osmoregulation: genetics and physiology.. Annu Rev Microbiol 1991; 45:569–606
    [Google Scholar]
  10. Dinnbier U., Limpinsel E., Schmid R., Bakker E. P. Transient accumulation of potassium glutamate and its replacement by trehalose during adaptation of growing cells of Escherichia coli K-12 to elevated sodium chloride concentrations. Arch Microbiol 1988; 150:348–357
    [Google Scholar]
  11. Frings E., Kunte H. J., Galinski E. A. Compatible solutes in representatives of the genera Brevibacterium and Corynebacterium : occurrence of tetrahydropyrimidines and glutamine.. FEMS Microbiol Lett 1993; 109:25–32
    [Google Scholar]
  12. Galinski E. A. Osmoadaptation in bacteria.. Adv Microb Physio 1995; 37:273–328
    [Google Scholar]
  13. Galinski E. A., Herzog R. M. The role of trehalose as a substitute for nitrogen-containing compatible solutes (Ectothiorhodospira halochloris) . Arch Microbiol 1990; 153:607–613
    [Google Scholar]
  14. Galinski E. A., Trüper H. G. Microbial behaviour in saltstressed ecosystems.. FEMS Microbiol Rev 1994; 15:95–108
    [Google Scholar]
  15. Gardan R., Rapoport G., Débarbouillé M. Expression of the rocDEF operon involved in arginine catabolism in Bacillus subtilis . J Mol Biol 1995; 249:843–856
    [Google Scholar]
  16. Gordia S., Gutierrez C. Growth-phase-dependent expression of the osmotically inducible gene osmC of Escherichia coli K-12. . Mol Microbiol 1996; 19:729–736
    [Google Scholar]
  17. Gouesbet G., Trautwetter A., Bonnassie S., Wu L. F., Blanco C. Characterization of the Erwinia chrysanthemi osmoprotectant transporter gene ousA . J Bacteriol 1996; 178:447–455
    [Google Scholar]
  18. Gowrishankar J, Manna D. How is osmotic regulation of transcription of the Escherichia coli proU operon achieved ?. Genetica 1996; 97:363–378
    [Google Scholar]
  19. Hecker M., Schumann W., Völker U. Heat-shock and general stress response in Bacillus subtilis .. Mol Microbiol 1996; 19:417–428
    [Google Scholar]
  20. Imhoff J. F., Trüper H. G. Ectothiorhodospira halochloris sp. nov., a new extremely halophilic phototrophic bacterium containing bacteriochlorophyll b.. Arch Microbiol 1977; 114:115–121
    [Google Scholar]
  21. Inoue Y., Kuramitsu S., Inoue K., Kagamiyama H., Hiromi K., Tanase S., Morino Y. Substitution of a lysyl residue for arginine 386 of Escherichia coli aspartate aminotransferase.. J Biol Chem 1989; 264:9673–9681
    [Google Scholar]
  22. Jebbar M., Talibart R., Gloux K., Bernard T., Blanco C. Osmoprotection of Escherichia coli by ectoine : uptake and accumulation characteristics.. J Bacteriol 1992; 174:5027–5035
    [Google Scholar]
  23. Kempf B., Bremer E. OpuA, an osmotically regulated binding protein-dependent transport system for the osmoprotectant glycine betaine in Bacillus subtilis .. J Biol Chem 1995; 270:16701–16713
    [Google Scholar]
  24. Kunte H. J., Galinski E. A., Galinski E. A., Trüper H. G. A modified FMOC-method for the detection of amino acid-type osmolytes and tetrahydropyrimidines (ectoines). . J Microbiol Methods 1993; 17:129–136
    [Google Scholar]
  25. Lamark T., Røkenes T. P., McDougal J., Strøm A. R. The complex bet promoters of Escherichia coli : regulation by oxygen (ArcA), choline (BetI), and osmotic stress.. J Bacteriol 1996; 178:1655–1662
    [Google Scholar]
  26. Larsen P. I., Sydne L. K., Landfald B., Strøm A. R. Osmoregulation in Escherichia coli by accumulation of organic osmolytes : betaines, glutamic acid, and trehalose.. Arch Microbiol 1987; 147:1–7
    [Google Scholar]
  27. Lucht J. M., Bremer E. Adaptation of Escherichia coli to high osmolarity environments : osmoregulation of the highaffinity glycine betaine transport system ProU.. FEMS Microbiol Rev 1994; 14:3–20
    [Google Scholar]
  28. Manna D., Gowrishankar J. Evidence for involvement of proteins HU and RpoS in transcription of the osmoresponsive proU operon in Escherichia coli . J Bacteriol 1994; 176:5378–5384
    [Google Scholar]
  29. Mehta P. K., Hale T. I., Christen P. Aminotransferases : demonstration of homology and division into evolutionary subgroups.. Eur J Biochem 1993; 214:549–561
    [Google Scholar]
  30. Mellies J., Brems R., Villarejo m. The Escherichia coli proU promoter element and its contribution to osmotically signaled transcription activation.. J Bacteriol 1994; 176:3638–3645
    [Google Scholar]
  31. Mellies J., Wise A., Villarejo M. Two different Escherichia coli proP promoters respond to osmotic and growth phase signals.. J Bacteriol 1995; 177:144–151
    [Google Scholar]
  32. Miller J. H. A Short Course in Bacterial Genetics: a Laboratory Manual and Handbook for Escherichia coli and Related Bacteria. Cold Spring Harbor Laboratory Cold Spring Harbor, NY:
    [Google Scholar]
  33. Min-Yu L., Ono H., Takano M. Gene cloning of ectoine synthase from Halomonas sp. Annu Rep Int Center Cooperative Res Biotechnol ,. Jpn 1993; 16:193–200
    [Google Scholar]
  34. O’Reilly M., Devine K. M. Sequence and analysis of the citrulline biosynthetic operon argC-F from Bacillus subtilis . . Microbiology 1994; 140:1023–1025
    [Google Scholar]
  35. Peters P., Galinski E. A., Trüper H. G. The biosynthesis of ectoine.. FEMS Microbiol Lett 1990; 71:157–162
    [Google Scholar]
  36. Severin J., Wohlfarth A., Galinsk E. A. The predominant role of recently discovered tetrahydropyrimidines for the osmoadaptation of halophilic eubacteria.. J Gen Microbiol 1992; 138:1629–1638
    [Google Scholar]
  37. Strøm A. R., Kaasen I. Trehalose metabolism in Escherichia coli : stress protection and stress regulation of gene expression.. Mol Microbiol 1993; 8:205–210
    [Google Scholar]
  38. Takeshita S., Sato M., Toba M., Masahashi W., HashimotoGotoh T. High-copy-number and low-copy-number plasmid vectors for lacZ alpha-complementation and chloramphenicol- or kanamycin-resistance selection. . Gene 1987; 61:63–74
    [Google Scholar]
  39. Tao T., Yasuda N., Ono H., Shinmyo A., Takano M. Purification and characterization of 2,4-diaminobutyric acid transaminase from Halomonas sp. Annu Rep lnt Center Cooperative Res Biotechnol ,. Jpn 1992; 15:187–199
    [Google Scholar]
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