Matthews R G, Daubner S C
Adv Enzyme Regul. 1982;20:123-31. doi: 10.1016/0065-2571(82)90012-7.
Methylenetetrahydrofolate reductase catalyzes the reduction of methylenetetrahydrofolate to methyltetrahydrofolate. This reaction commits one carbon units to the pathways of adenosylmethionine-dependent methylation in mammalian cells. We have purified the pig liver enzyme to homogeneity and shown that it contains FAD as a non-covalently bound prosthetic group. Methylenetetrahydrofolate is not only a substrate for the reductase, but also for thymidylate synthase and for methylenetetrahydrofolate dehydrogenase. The latter reaction leads to utilization of one carbon units in de novo purine biosynthesis. A priori, one might expect that methylenetetrahydrofolate reductase activity would be modulated by cellular requirements for de novo biosynthesis of purines and pyrimidines, as well as by cellular levels of adenosylmethionine. Methylenetetrahydrofolate reductase is inhibited by dihydrofolate and its polyglutamate analogues. The Ki is 6.5 microM for dihydrofolate and decreases with each additional glutamyl residue to a minimum value of 0.013 microM for dihydropteroylhexaglutamate. The I50 for dihydropteroylhexaglutamate inhibition of reductase activity in the presence of 0.5 microM methylenetetrahydropteroylhexaglutamate is 0.07 microM. We propose that stimulation of thymidylate synthase activity (as in the replicating cell) may lead to elevations in the steady state levels of cellular dihydrofolate derivatives and to resultant inhibition of methylenetetrahydrofolate reductase activity. Thus methylenetetrahydrofolate derivatives would be spared for purine and pyrimidine biosynthesis. We have also examined the inhibition of methylenetetrahydrofolate reductase by adenosylmethionine, which serves as an allosteric effector of the enzymatic activity. Adenosylmethionine induces a slow transition in the enzyme, and leads to the inhibition of NADPH-menadione, NADPH-methylenetetrahydrofolate and methyltetrahydrofolate-menadione oxido-reductase activities.
亚甲基四氢叶酸还原酶催化亚甲基四氢叶酸还原为甲基四氢叶酸。该反应使一碳单位进入哺乳动物细胞中依赖腺苷甲硫氨酸的甲基化途径。我们已将猪肝酶纯化至同质，并表明它含有FAD作为非共价结合的辅基。亚甲基四氢叶酸不仅是还原酶的底物，也是胸苷酸合成酶和亚甲基四氢叶酸脱氢酶的底物。后一反应导致一碳单位在嘌呤从头生物合成中的利用。据推测，亚甲基四氢叶酸还原酶活性可能受细胞对嘌呤和嘧啶从头生物合成的需求以及细胞内腺苷甲硫氨酸水平的调节。亚甲基四氢叶酸还原酶受二氢叶酸及其多聚谷氨酸类似物的抑制。二氢叶酸的Ki为6.5微摩尔，随着每个额外的谷氨酰残基，其值降低，对于二氢蝶酰六谷氨酸，其最小值为0.013微摩尔。在存在0.5微摩尔亚甲基四氢蝶酰六谷氨酸的情况下，二氢蝶酰六谷氨酸对还原酶活性的I50为0.07微摩尔。我们提出，胸苷酸合成酶活性（如在复制细胞中）的刺激可能导致细胞二氢叶酸衍生物稳态水平的升高，并导致亚甲基四氢叶酸还原酶活性的抑制。因此，亚甲基四氢叶酸衍生物将被节省用于嘌呤和嘧啶的生物合成。我们还研究了腺苷甲硫氨酸对亚甲基四氢叶酸还原酶的抑制作用，腺苷甲硫氨酸作为酶活性的变构效应物。腺苷甲硫氨酸诱导酶的缓慢转变，并导致对NADPH-甲萘醌、NADPH-亚甲基四氢叶酸和甲基四氢叶酸-甲萘醌氧化还原酶活性的抑制。
