Fu T F, Rife J P, Schirch V
Department of Biochemistry, Institute for Structural Biology and Drug Discovery, 800 East Leigh Street, Suite 212, Richmond, Virginia 23219, USA.
Arch Biochem Biophys. 2001 Sep 1;393(1):42-50. doi: 10.1006/abbi.2001.2471.
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-(13)C]serine, [2-(13)C]glycine, and [(13)C]formate. (13)C 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 (13)C label in purines was determined from the splitting patterns of the (1)H 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-(13)C]Glycine is not a donor of one-carbons groups, confirming that MCF-7 cells lack a functional glycine cleavage system.
通过观察叶酸介导的L-[3-(13)C]丝氨酸、[2-(13)C]甘氨酸和[(13)C]甲酸的一碳代谢，研究了胞质和线粒体丝氨酸羟甲基转移酶在为MCF-7细胞中嘌呤和胸苷酸生物合成提供一碳基团方面的作用。利用(13)C NMR追踪标记物掺入嘌呤的C2和C8以及胸苷酸C5上连接的甲基。根据腺嘌呤C2和C8以及鸟嘌呤C8的(1)H NMR谱的分裂模式确定嘌呤中(13)C标记的富集百分比。结果表明，甲酸是10-甲酰四氢叶酸中用于嘌呤生物合成的一碳基团以及5,10-亚甲基四氢叶酸中用于胸苷酸生物合成的一碳基团在胞质中的主要前体。甲酸由丝氨酸的C3在线粒体中形成。胞质丝氨酸羟甲基转移酶将丝氨酸裂解为甘氨酸和5,10-亚甲基四氢叶酸似乎不是嘌呤或胸苷酸生物合成一碳基团的主要来源。丝氨酸的C3约占一碳池的95%，这表明一碳基团的其他来源仅代表次要途径。[2-(13)C]甘氨酸不是一碳基团的供体，证实MCF-7细胞缺乏功能性甘氨酸裂解系统。
