1887

Abstract

In the yeast , the first committed step of the lysine biosynthetic pathway is catalysed by two homocitrate synthases encoded by and . We undertook a study of the duplicate homocitrate synthases to analyse whether their retention and presumable specialization have affected the efficiency of lysine biosynthesis in yeast. Our results show that during growth on ethanol, homocitrate is mainly synthesized through Lys21p, while under fermentative metabolism, Lys20p and Lys21p play redundant roles. Furthermore, results presented in this paper indicate that, in contrast to that which had been found for Lys20p, lysine is a strong allosteric inhibitor of Lys21p ( 0.053 mM), which, in addition, induces positive co-operativity for -ketoglutarate (-KG) binding. Differential lysine inhibition and modulation by -KG of the two isozymes, and the regulation of the intracellular amount of the two isoforms, give rise to an exquisite regulatory system, which balances the rate at which -KG is diverted to lysine biosynthesis or to other metabolic pathways. It can thus be concluded that retention and further biochemical specialization of the - and -encoded enzymes with partially overlapping roles contributed to the acquisition of facultative metabolism.

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2008-06-01
2024-04-27
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References

  1. Andi B., West A. H., Cook P. F. 2004a; Kinetic mechanism of histidine-tagged homocitrate synthase from Saccharomyces cerevisiae . Biochemistry 43:11790–11795
    [Google Scholar]
  2. Andi B., West A. H., Cook P. F. 2004b; Stabilization and characterization of histidine-tagged homocitrate synthase from Saccharomyces cerevisiae . Arch Biochem Biophys 421:243–254
    [Google Scholar]
  3. Andi B., West A. H., Cook P. F. 2005; Regulatory mechanism of histidine-tagged homocitrate synthase from Saccharomyces cerevisiae . I. Kinetic studies. J Biol Chem 280:31624–31632
    [Google Scholar]
  4. Avendaño A., Riego L., DeLuna A., Aranda C., Romero G., Ishida C., Vázquez-Acevedo M., Rodarte B., Recillas-Targa F. other authors 2005; Swi/SNF–GCN5–dependent chromatin remodelling determines induced expression of GDH3 , one of the paralogous genes responsible for ammonium assimilation and glutamate biosynthesis in Saccharomyces cerevisiae . Mol Microbiol 57:291–305
    [Google Scholar]
  5. Betterton H., Fjellstedt T., Matsuda M., Ogur M., Tate R. 1968; Localization of the homocitrate pathway. Biochim Biophys Acta 170:459–461
    [Google Scholar]
  6. Chen J. C. Y., Powers T. 2006; Coordinate regulation of multiple and distinct biosynthetic pathways by TOR and PKA kinases in S. cerevisiae . Curr Genet 49:281–293
    [Google Scholar]
  7. Chen S., Brockenbrough J. S., Dove J. E., Aris J. P. 1997; Homocitrate synthase is located in the nucleus in the yeast Saccharomyces cerevisiae . J Biol Chem 272:10839–10846
    [Google Scholar]
  8. Christianson T. W., Sikorski R. S., Dante M., Shero J. H., Hieter P. 1992; Multifunctional yeast high-copy-number shuttle vectors. Gene 110:119–122
    [Google Scholar]
  9. DeLuna A., Avendaño A., Riego L., González A. 2001; NADP-glutamate dehydrogenase isoenzymes of Saccharomyces cerevisiae . Purification, kinetic properties, and physiological roles. J Biol Chem 276:43775–43783
    [Google Scholar]
  10. Eisler H., Frohlich K., Heidenreich E. 2004; Starvation for an essential amino acid induces apoptosis and oxidative stress in yeast. Exp Cell Res 300:345–353
    [Google Scholar]
  11. Feller A., Dubois E., Ramos F., Pierard A. 1994; Repression of the genes for lysine biosynthesis in Saccharomyces cerevisiae is caused by limitation of Lys14-dependent transcriptional activation. Mol Cell Biol 14:6411–6418
    [Google Scholar]
  12. Feller A., Ramos F., Pierard A., Dubois E. 1999; In Saccharomyces cerevisiae , feedback inhibition of homocitrate synthase isoenzymes by lysine modulates the activation of LYS gene expression by Lys14p. Eur J Biochem 261:163–170
    [Google Scholar]
  13. Force A., Lynch M., Pickett F. B., Amores A., Yan Y. L., Postlethwait J. 1999; Preservation of duplicate genes by complementary, degenerative mutations. Genetics 151:1531–1545
    [Google Scholar]
  14. Gaillardin C. M., Poirier L., Heslot H. 1976; A kinetic study of homocitrate synthetase activity in the yeast Saccharomycopsis lipolytica . Biochim Biophys Acta 422:390–406
    [Google Scholar]
  15. González B., Francois J., Renaud M. 1997; A rapid and reliable method for metabolite extraction in yeast using boiling buffered ethanol. Yeast 13:1347–1356
    [Google Scholar]
  16. Ito H., Fukuda Y., Murata K., Kimura A. 1983; Transformation of intact yeast cells treated with alkali cations. J Bacteriol 153:163–168
    [Google Scholar]
  17. Kosuge T., Hoshino T. 1998; Lysine is synthesized through the α -aminoadipate pathway in Thermus thermophilus . FEMS Microbiol Lett 169:361–367
    [Google Scholar]
  18. Kwon E. S., Jeong J. H., Roe J. H. 2006; Inactivation of homocitrate synthase causes lysine auxotrophy in copper/zinc-containing superoxide dismutase-deficient yeast Schizosaccharomyces pombe . J Biol Chem 281:1345–1351
    [Google Scholar]
  19. Lowry O. H., Rosebrough N. J., Farr A. L., Randall R. J. 1951; Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275
    [Google Scholar]
  20. Lynch M., Force A. 2000; The probability of duplicate gene preservation by subfunctionalization. Genetics 154:459–473
    [Google Scholar]
  21. Merico A., Sulo P., Piskur J., Compagno C. 2007; Fermentative lifestyle in yeasts belonging to the Saccharomyces complex. FEBS J 274:976–989
    [Google Scholar]
  22. Monod J., Wyman J., Changeux J. P. 1965; On the nature of allosteric transitions: a plausible model. J Mol Biol 12:88–118
    [Google Scholar]
  23. Natarajan K., Meyer M. R., Jackson B. M., Slade D., Roberts C., Hinnebusch A. G., Marton M. J. 2001; Transcriptional profiling shows that Gcn4p is a master regulator of gene expression during amino acid starvation in yeast. Mol Cell Biol 21:4347–4368
    [Google Scholar]
  24. Nishida H., Nishiyama M. 2000; What is characteristic of fungal lysine synthesis through the α -aminoadipate pathway?. J Mol Evol 51:299–302
    [Google Scholar]
  25. Nishida H., Nishiyama M., Kobashi N., Kosuge T., Hoshino T., Yamane H. 1999; A prokaryotic gene cluster involved in synthesis of lysine through the amino adipate pathway: a key to the evolution of amino acid biosynthesis. Genome Res 9:1175–1183
    [Google Scholar]
  26. Piskur J. 2001; Origin of the duplicated regions in the yeast genomes. Trends Genet 17:302–303
    [Google Scholar]
  27. Ramos F., Wiame J. M. 1985; Mutation affecting the specific regulatory control of lysine biosynthetic enzymes in Saccharomyces cerevisiae . Mol Gen Genet 200:291–294
    [Google Scholar]
  28. Ramos F., Dubois E., Pierard A. 1988; Control of enzyme synthesis in the lysine biosynthetic pathway of Saccharomyces cerevisiae . Evidence for a regulatory role of gene LYS14 . Eur J Biochem 171:171–176
    [Google Scholar]
  29. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  30. Segel I. H. 1975 Enzyme Kinetics: Behavior and Analysis of Rapid Equilibrium and Steady State Enzyme Systems pp 346–462 New York: Wiley;
    [Google Scholar]
  31. Sikorski R. S., Hieter P. 1989; A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae . Genetics 122:19–27
    [Google Scholar]
  32. Stephen D. W. S., Jamieson D. J. 1997; Amino acid-dependent regulation of the Saccharomyces cerevisiae GSH1 gene by hydrogen peroxide. Mol Microbiol 23:203–210
    [Google Scholar]
  33. Struhl K., Davis R. W. 1981; Transcription of the his3 gene region in Saccharomyces cerevisiae . J Mol Biol 152:535–552
    [Google Scholar]
  34. Takagi H., Iwamoto F., Nakamori S. 1997; Isolation of freeze-tolerant laboratory strains of Saccharomyces cerevisiae from proline-analogue-resistant mutants. Appl Microbiol Biotechnol 47:405–411
    [Google Scholar]
  35. Tate J. J., Feller A., Dubois E., Cooper T. G. 2006; Saccharomyces cerevisiae Sit4 phosphatase is active irrespective of the nitrogen source provided, and Gln3 phosphorylation levels become nitrogen source-responsive in a sit4 -deleted strain. J Biol Chem 281:37980–37992
    [Google Scholar]
  36. Towbin H., Staehelin T., Gordon J. 1979; Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76:4350–4354
    [Google Scholar]
  37. Valenzuela L., Aranda C., González A. 2001; TOR modulates GCN4-dependent expression of genes turned on by nitrogen limitation. J Bacteriol 183:2331–2334
    [Google Scholar]
  38. Watson T. G. 1976; Amino-acid pool composition of Saccharomyces cerevisiae as a function of growth rate and amino-acid nitrogen source. J Gen Microbiol 96:263–268
    [Google Scholar]
  39. Wolfe K. H., Shields D. C. 1997; Molecular evidence for an ancient duplication of the entire yeast genome. Nature 387:708–713
    [Google Scholar]
  40. Xu H., Andi B., Qian J., West A. H., Cook P. F. 2006; The α -aminoadipate pathway for lysine biosynthesis in fungi. Cell Biochem Biophys 46:43–64
    [Google Scholar]
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