The Role of Serine Hydroxymethyltransferase Isozymes in One-Carbon Metabolism in MCF-7 Cells as Determined by 13C NMR

Abstract

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-[3-13C]serine, [2-13C]glycine, and [13C]formate. 13C NMR 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 13C label in purines was determined from the splitting patterns of the 1H 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. [2-13C]Glycine is not a donor of one-carbons groups, confirming that MCF-7 cells lack a functional glycine cleavage system.