The roles of serine hydroxymethyltransferase and methylenetetrahydrofolate reductase isozymes in one-carbon metabolism.

摘要

Serine hydroxymethyltransferase (SHMT) catalyzes the conversion of serine to 5,10-methylene-THF and glycine. In Saccharomyces cerevisiae, two genes have been isolated and sequenced, encoding for a cytoplasmic (SHM2) and a mitochondrial (SHM1) isozyme (McNeil et al., 1994). In the same study, deletion of the two loci resulted in no growth requirement. In my work, ser1{dollar}sp-{dollar} strains containing deletions at either one or both loci were generated. The identity of the strains was confirmed by Southern analysis, subfractionation experiments, and activity assays. Growth studies using serine, glycine, and formate as alternative one-carbon sources suggest different roles for the two isozymes. The mitochondrial isozyme is thought to play a major role in providing serine, whereas the cytoplasmic isozyme mainly provides one-carbon units and glycine. Evidence supporting the presence of an additional source of glycine (McNeil et al., 1994) is also provided by the growth studies. In order to quantitate the contributions of the two isozymes in the provision of one-carbon units or glycine, {dollar}sp{lcub}13{rcub}{dollar}C labeling experiments and NMR were used. Competition experiments with unlabeled serine suggest that both isozymes provide one-carbon units for purine biosynthesis. However, the cytoplasmic isozyme contributes more to the glycine pool used for purine biosynthesis than the mitochondrial isozyme.; 5,10-methylene-THF reductase generates the methyl group of methionine by irreversibly reducing 5,10-methylene-THF to 5-methyl-THF. In mammals, this reaction is the only pathway for the de novo synthesis of the methyl group of methionine (Matthews, 1984). Based on homology comparisons with MTHFR sequences from other organisms, two genes encoding for MTHFR in S. cerevisiae were isolated and sequenced (designated MTR1 and MTR2). The 3{dollar}spprime{dollar}-end of MTR2 has not yet been isolated. Homology comparisons suggest that MTR1 has a similar two-domain structure as the human MTHFR and an S-adenosylmethionine binding site. Deletion of the MTR1 locus results in no growth requirement in this strain, suggesting that the product of MTR2 may also be a functional reductase. These studies highlight the different metabolic roles of the isozymes of these two important activities in folate metabolism.