The role of serine hydroxymethyltransferase isozymes in one-carbonmetabolism in MCF-7 cells as determined by C-13 NMR

摘要

The role of cytosolic and mitochondrial serine hydroxymethyltransferase in supplying one-carbon groups for purine and thymidylate biosynthesis in MCF-7 cells was investigated by observing folate-mediated one-carbon metabolism of L-[3-C-13]serine, [2-C-13]glycine, and [C-13]formate. C-13 NAM was used to follow the incorporation of label into carbons 2 and 8 of purines and the methyl group attached to carbon 5 of thymidylate. The percentage enrichment of the C-13 label in purines was determined from the splitting patterns of the H-1 NMR spectra of C2 and C8 of adenine and C8 of guanine. The results show that formate is the major precursor in the cytosol of the one-carbon group in 10-formyltetrahydrofolate, which is used in purine biosynthesis, and the one-carbon group in 5,10-methylenetetrahydrofolate, which is used in thymidylate biosynthesis. Formate is formed in the mitochondria from carbon 3 of serine. The cleavage of serine to glycine and 5,10-methylenetetrahydrofolate by cytosolic serine hydroxymethyltransferase does not appear to be a major source of one-carbon groups for either purine or thymidylate biosynthesis. Carbon 3 of serine accounts for about 95% of the one-carbon pool, suggesting that other sources of one-carbon groups represent only minor pathways. [2-C-13]Glycine is not a donor of one-carbons groups, confirming that MCF-7 cells lack a functional glycine cleavage system.