STRUCTURAL FEATURES OF 5,10-DIDEAZA-5,6,7,8-TETRAHYDROFOLATE THAT DETERMINE INHIBITION OF MAMMALIAN GLYCINAMIDE RIBONUCLEOTIDE FORMYLTRANSFERASE
STRUCTURAL FEATURES OF 5,10-DIDEAZA-5,6,7,8-TETRAHYDROFOLATE THAT DETERMINE INHIBITION OF MAMMALIAN GLYCINAMIDE RIBONUCLEOTIDE FORMYLTRANSFERASE
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DOI：
10.1021/bi00221a037
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发表时间：
1991-02-19
期刊：
影响因子：
2.9
通讯作者：
MORAN, RG
中科院分区：
文献类型：
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作者：
BALDWIN, SW;TSE, A;MORAN, RG
我们研究了 5,10-二脱氮杂-5,6,7,8-四氢叶酸 (DDATHF) 的结构特征，确定了该化合物作为从小鼠 L 1210 细胞纯化的甘氨酰胺核糖核苷酸甲酰基转移酶 (GARFT) 抑制剂的活性。 5-脱氮四氢叶酸是与 DDATHF 一样好的 GARFT 抑制剂，表明四氢叶酸 5 位碳等排取代氮足以抑制 GARFT。 5,10-二脱氮叶酸、5,8,10-三氮杂四氢叶酸和 2-脱氨基-5,10-二脱氮四氢叶酸是 GARFT 的弱抑制剂，表明还原的吡啶并嘧啶环、N-8 和 DDATHF 的 2-氨基分别在四氢叶酸类似物与该酶的结合中发挥重要作用。 DDATHF 类似物（其中苯环被环己基环或亚甲基取代）保留了作为抑制剂的活性。 5,10-二脱氮四氢高叶酸作为 GARFT 抑制剂的效力比 DDATHF 强约 6 倍，但 5,10-二脱氮四氢去甲叶酸的活性约为 DDATHF 的五分之一。 DDATHF 的类似物，其中谷氨酸侧链被天冬氨酸取代（天冬氨酸不是多聚谷氨酸化的底物，仅具有弱细胞毒性），作为纯化 GARFT 的抑制剂，其活性与 DDATHF 等效。 令人惊讶的是，作为 GARFT 抑制剂，5,10-二脱氮四氢蝶呤酸的活性与 DDATHF 大致相同，这表明 DDATHF 侧链中的谷氨酸在这种配体-酶相互作用中不起作用。 DDATHF 的聚谷氨酸衍生物与 GARFT 的结合比 DDATHF 本身的结合紧密 100 倍；长链聚谷氨酸盐在实验条件下符合 Goldstein 的 B 区行为，预计在体内处于 C 区，即化学计量抑制。 我们得出结论，四氢叶酸类似物 5 位碳的存在足以抑制 GARFT，N-8 和 2-氨基可能通过氢键参与 DDATHF 与 GARFT 的结合，并且 DDATHF 类似物的苯环和氨基酸侧链的结构不是这些化合物的单谷氨酸盐形式抑制 GARFT 的主要决定因素。 我们还得出结论，多谷氨酸化在 DDATHF 的有效细胞毒性中发挥着重要作用。
We have investigated the structural features of 5,10-dideaza-5,6,7,8-tetrahydrofolate (DDATHF) that determine the activity of this compound as an inhibitor of glycinamide ribonucleotide formyltransferase (GARFT) purified from mouse L 1210 cells. 5-Deazatetrahydrofolate was as good an inhibitor of GARFT as DDATHF, indicating that isosteric replacement of nitrogen by carbon at the 5-position of tetrahydrofolate is sufficient for inhibition of GARFT. 5,10-Dideazafolic acid, 5,8,10-trideazatetrahydrofolate, and 2-desamino-5,10-dideazatetrahydrofolate were poor inhibitors of GARFT, indicating that a reduced pyridopyrimidine ring, N-8, and the 2-amino group of DDATHF, respectively, play an important role in the binding of tetrahydrofolate analogues to this enzyme. DDATHF analogues in which the phenyl ring was replaced either by a cyclohexyl ring or by methylene groups retained activity as inhibitors. 5,10-Dideazatetrahydrohomofolate was about 6 times more potent as an inhibitor of GARFT than DDATHF, but 5,10-dideazatetrahydronorfolate had about one-fifth of the activity of DDATHF. An analogue of DDATHF in which the glutamic acid side chain was replaced by aspartic acid (which was not a substrate for polyglutamation and was only weakly cytotoxic) was equiactive with DDATHF as an inhibitor of purified GARFT. Surprisingly, 5,10-dideazatetrahydropteroic acid was about as active as DDATHF as an inhibitor of GARFT, an indication that the glutamic acid in the side chain of DDATHF does not play a role in this ligand-enzyme interaction. The polyglutamate derivatives of DDATHF bound up to 100 times tighter to GARFT than DDATHF itself; longer chain polyglutamates conformed to Goldstein's zone B behavior under experimental conditions and were projected to be in zone C, i.e., stoichiometric inhibition, in vivo. We conclude that the presence of carbon at the 5-position of tetrahydrofolate analogues is sufficient for inhibition of GARFT, that N-8 and the 2-amino group are involved in binding of DDATHF to GARFT, probably through hydrogen bonds, and that the structures of the phenyl ring and amino acid side chain of DDATHF analogues are not primary determinants of GARFT inhibition by monoglutamate forms of these compounds. We also conclude that polyglutamation plays a major role in the potent cytotoxicity of DDATHF.