ROLE OF MITOCHONDRIAL AND CYTOPLASMIC SERINE HYDROXYMETHYLTRANSFERASEISOZYMES IN DE-NOVO PURINE SYNTHESIS IN SACCHAROMYCES-CEREVISIAE

One-carbon units are essential to a variety of anabolic processes whic h yield necessary cellular components including purines, pyrimidines, amino acids, and lipids, Serine hydroxymethyltransferase (SHMT) is the major provider of one-carbon units in the cell, The other product of this reaction is glycine, Both of these metabolites are required In de novo purine biosynthesis. In Saccharomyces cerevisiae, mitochondrial and cytoplasmic SHMT isozymes are encoded by distinct nuclear genes (S HM1 and SHM2). Molecular genetic analyses have begun to define the rol es of these two isozymes in folate-mediated one-carbon metabolism [McN eil, J. B., et al. (1996) Genetics 142, 371-381]. In our study, the SH M1 and SHM2 genes were disrupted singly and in combination to investig ate the contributions of the two SHMT isozymes to tile production of g lycine and one-carbon units required in purine biosynthesis. Cell subf ractionation experiments indicated that while only 5% of total activit y was localized in the mitochondria, the specific activity in that com partment was much higher than in the cytoplasm. Growth and C-13 NMR ex periments indicate that the two isozymes function in different directi ons, depending on the nutritional conditions of the cell. When yeast w as grown on serine as the primary one-carbon source, the cytoplasmic i sozyme was the main provider of glycine and one-carbon groups for puri ne synthesis. When grown on glycine, the mitochondrial SHMT was the pr edominant isozyme catalyzing the synthesis of serine from glycine and one-carbon units. However, when both serine and glycine were present, the mitochondrial SHMT made a significant contribution of one-carbon u nits, but not glycine, for purine synthesis, Finally, NMR data are pre sented that suggest the existence of at least two sites of de novo pur ine biosynthesis in growing yeast cells, each being fed by distinct po ols of precursors.