Mejia N R, Rios-Orlandi E M, MacKenzie R E
J Biol Chem. 1986 Jul 15;261(20):9509-13.
NAD-dependent methylenetetrahydrofolate dehydrogenase is expressed in transformed or established mammalian cell lines in vitro but only in the developmental tissues of normal adult animals (Mejia, N. R. and MacKenzie, R. E. (1985) J. Biol. Chem. 260, 14616-14620). The enzyme, which contains methenyltetrahydrofolate cyclohydrolase activity as well, has been purified 6000-fold from Ehrlich ascites tumor cells. The preparation is homogeneous by sodium dodecyl sulfate gel electrophoresis (Mr = 34,000), and results from cross-linking with bis(sulfosuccinimidyl)suberate are consistent with a dimeric structure (Mr = 68,000) for the native bifunctional enzyme. The dehydrogenase is specific for NAD and requires both a divalent cation, Mg2+ or Mn2+, for activity and as well is stimulated by inorganic phosphate. When compared to the usual NADP-dependent methylenetetrahydrofolate dehydrogenase from mouse liver, the NAD-dependent dehydrogenase activity of the murine tumor enzyme shows a greater affinity for the polyglutamate forms of folate.
NAD 依赖性亚甲基四氢叶酸脱氢酶在体外转化的或已建立的哺乳动物细胞系中表达，但仅在正常成年动物的发育组织中表达（梅希亚，N.R.和麦肯齐，R.E.（1985 年）《生物化学杂志》260，14616 - 14620）。该酶还具有亚甲基四氢叶酸环水解酶活性，已从艾氏腹水瘤细胞中纯化了 6000 倍。通过十二烷基硫酸钠凝胶电泳（Mr = 34,000），该制剂是均一的，并且用辛二酸双（磺基琥珀酰亚胺酯）交联的结果与天然双功能酶的二聚体结构（Mr = 68,000）一致。该脱氢酶对 NAD 具有特异性，活性需要二价阳离子 Mg2 + 或 Mn2 +，并且也受到无机磷酸盐的刺激。与来自小鼠肝脏的通常的 NADP 依赖性亚甲基四氢叶酸脱氢酶相比，鼠肿瘤酶的 NAD 依赖性脱氢酶活性对叶酸的多聚谷氨酸形式表现出更高的亲和力。
