1887

Abstract

Methionine synthase (EC2.1.1.14) catalyses the final step in methionine synthesis, i.e. methylation of homocysteine. A search of the genomic database revealed a gene designated SPAC9.09, encoding a protein with significant homology to methionine synthase. Disruption of SPAC9.09 caused methionine auxotrophy, and thus the gene was identified as a methionine synthase and designated . The mutant was found to exhibit a remarkable growth defect in the absence of adenine even in medium supplemented with methionine. This phenotype was not observed in other methionine auxotrophs. In the budding yeast , which has been reported to utilize homocysteine in cysteine synthesis, lack of a functional methionine synthase did not cause a requirement for adenine. The introduction of genes from constituting the cystathionine pathway ( and ) into Δ cells restored growth in the absence of adenine. HPLC analysis showed that total homocysteine content in Δ cells was higher than in other methionine auxotrophs and that introduction of the cystathionine pathway decreased total homocysteine levels. These data demonstrate that accumulation of homocysteine causes a defect in purine biosynthesis in the mutant.

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2006-02-01
2020-04-01
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References

  1. Aguilar B, Rojas J. C, Collados M. T. 2004; Metabolism of homocysteine and its relationship with cardiovascular disease. J Thromb Thrombolysis18:75–87[CrossRef]
    [Google Scholar]
  2. Alfa C, Fantes P, Hyams J, Mcleod M, Warbrick E. 1993; Experiments with Fission Yeast. A Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  3. Allen E. H, Hussey G. G. 1971; Inhibition of the growth of Helminthosporium carbonum by l-cysteine. Can J Microbiol17:101–103[CrossRef]
    [Google Scholar]
  4. Basi G, Schmid E, Maundrell K. 1993; TATA box mutations in the Schizosaccharomyces pombe nmt1 promoter affect transcription efficiency but not the transcription start point or thiamine repressibility. Gene123:131–136[CrossRef]
    [Google Scholar]
  5. Brzywczy J, Sienko M, Kucharska A, Paszewski A. 2002; Sulphur amino acid synthesis in Schizosaccharomyces pombe represents a specific variant of sulphur metabolism in fungi. Yeast19:29–35[CrossRef]
    [Google Scholar]
  6. Chaudhuri B, Ingavale S, Bachhawat A. K. 1996; apd1 [sup]+[/sup], a gene required for red pigment formation in ade6 mutants of Schizosaccharomyces pombe , encodes an enzyme required for glutathione biosynthesis: a role for glutathione and a glutathione-conjugate pump. Genetics145:75–83
    [Google Scholar]
  7. Fujita Y, Takegawa K. 2004; Characterization of two genes encoding putative cysteine synthase required for cysteine biosynthesis in Schizosaccharomyces pombe . Biosci Biotechnol Biochem68:306–311[CrossRef]
    [Google Scholar]
  8. Fujita Y, Giga-Hama Y, Takegawa K. 2005; Development of a genetic transformation system using new selectable markers for fission yeast Schizosaccharomyces pombe . Yeast22:193–202[CrossRef]
    [Google Scholar]
  9. Grimm C, Kohil J, Murray J, Maundrell K. 1988; Genetic engineering of Schizosaccharomyces pombe : a system for gene disruption and replacement using the ura4 gene as a selectable marker. Mol Gen Genet215:81–86[CrossRef]
    [Google Scholar]
  10. Hatanaka H, Ariga N, Nagai J, Katsuki H. 1974; Accumulation of a sterol intermediate during reaction in the presence of homocysteine with cell-free extract of yeast. Biochem Biophys Res Commun60:787–793[CrossRef]
    [Google Scholar]
  11. Holmes W. B, Appling D. R. 2002; Cloning and characterization of methenyltetrahydrofolate synthetase from Saccharomyces cerevisiae . J Biol Chem277:20205–20213[CrossRef]
    [Google Scholar]
  12. Iwaki T, Takegawa K. 2004; A set of loxP marker cassettes for Cre-mediated multiple gene disruption in Schizosaccharomyces pombe . Biosci Biotechnol Biochem68:545–550[CrossRef]
    [Google Scholar]
  13. Kari C, Nagy Z, Kovacs P, Hernadi F. 1971; Mechanism of the growth inhibitory effect of cysteine on Escherichia coli . J Gen Microbiol68:349–356[CrossRef]
    [Google Scholar]
  14. Klein R. D, Favreau M. A. 1988; Transformation of Schwanniomyces occidentalis with an ADE2 gene cloned from S. occidentalis . J Bacteriol170:5572–5578
    [Google Scholar]
  15. Marzluf G. A. 1997; Molecular genetics of sulfur assimilation in filamentous fungi and yeast. Annu Rev Microbiol51:73–96[CrossRef]
    [Google Scholar]
  16. Maw G. A. 1961; Ability of S -methyl-l-cysteine to annul the inhibition of yeast growth by l-ethionine and by s -ethyl-l-cysteine. J Gen Microbiol25:441–449[CrossRef]
    [Google Scholar]
  17. Morita T, Takegawa K. 2004; A simple and efficient procedure for transformation of Schizosaccharomyces pombe . Yeast21:613–617[CrossRef]
    [Google Scholar]
  18. Parks L. W, Casey W. M. 1995; Physiological implications of sterol biosynthesis in yeast. Annu Rev Microbiol49:95–116[CrossRef]
    [Google Scholar]
  19. Paszewski A. 1993; Sulfur amino acid metabolism and its regulation in fungi: studies with Aspergillus nidulans . Acta Biochim Pol40:445–449
    [Google Scholar]
  20. Quere I, Habib A, Tobelem G, Maclouf J. 1995; Inhibition of cyclooxygenase activity in human endothelial cells by homocysteine. Adv Prostaglandin Thromboxane Leukot Res23:397–399
    [Google Scholar]
  21. Raposo B, Rodriguez C, Martinez-Gonzalez J, Badimon L. 2004; High levels of homocysteine inhibit lysyl oxidase (LOX) and downregulate LOX expression in vascular endothelial cells. Atherosclerosis177:1–8[CrossRef]
    [Google Scholar]
  22. Roman H. 1956; A system selective for mutations affecting the synthesis of adenine in yeast. Compt Rend Trav Lab Carlsberg Ser Physiol26:299–314
    [Google Scholar]
  23. Schweingruber A. M, Hilti N, Edenharter E, Schweingruber M. E. 1998; Methionine induces sexual development in the fission yeast Schizosaccharomyces pombe via an ste11 -dependent signalling pathway. J Bacteriol180:6338–6341
    [Google Scholar]
  24. Suga M, Hatakeyama T. 2001; High efficiency transformation of Schizosaccharomyces pombe pretreated with thiol compounds by electroporation. Yeast18:1015–1021[CrossRef]
    [Google Scholar]
  25. Thomas D, Surdin-Kerjan Y. 1997; Metabolism of sulfur amino acids in Saccharomyces cerevisiae . Microbiol Mol Biol Rev61:503–532
    [Google Scholar]
  26. Topal G, Brunet A, Millanvoye E, Boucher J. L, Rendu F, Devynck M. A, David-Dufilho M. 2004; Homocysteine induces oxidative stress by uncoupling of NO synthase activity through reduction of tetrahydrobiopterin. Free Radic Biol Med36:1532–1541[CrossRef]
    [Google Scholar]
  27. Wickerham L. J. 1946; A critical evaluation of the nitrogen assimilation tests commonly used in the classification of yeast. J Bacteriol52:293–301
    [Google Scholar]
  28. Zonneveld B. J, van der Zanden A. L. 1995; The red ade mutants of Kluyveromyces lactis and their classification by complementation with cloned ADE1 or ADE2 genes from Saccharomyces cerevisiae . Yeast11:823–827[CrossRef]
    [Google Scholar]
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