Mammalian folylpoly-gamma-glutamate synthetase. 3. Specificity for folate analogues.
Mammalian folylpoly-gamma-glutamate synthetase. 3. Specificity for folate analogues.
复制标题
哺乳动物叶酰聚-γ-谷氨酸合成酶。
DOI：
10.1021/bi00376a026
复制
发表时间：
1987
期刊：
影响因子：
2.9
通讯作者：
Shane,B
中科院分区：
文献类型：
--
作者：
George,S;Cichowicz,DJ;Shane,B
摘要：为了探索猪肝叶酰聚谷氨酸合成酶叶酸结合位点的特异性和催化条件，合成了多种叶酸类似物。叶酸的内部和末端谷氨酸部分的修饰引起对蛋白质的结合速率和/或亲和力的大幅下降。唯一的例外是谷氨酰胺，同型半胱氨酸，和鸟氨酸类似物，表明一个不太严格的特异性周围的碳谷氨酸。有人提出，初始叶酸bindingto酶涉及低亲和力的相互作用，在蝶呤和谷氨酸的网站和第一谷氨酸结合是内部残基相邻的苯甲酰基。当末端残基结合时，聚谷氨酸链通过谷氨酸位点的过程性运动和蛋白质中可能的构象变化将导致紧密结合，并将末端谷氨酸的羧基定位在催化的正确位置。施加在环出的内部谷氨酸残基上的空间限制和由不同蝶呤部分的结合施加的额外空间约束将预期影响蛋白质和/或末端谷氨酸的轻微构象变化，并且将解释随着聚谷氨酸链长度的增加，开启速率和催化速率的降低，以及一碳取代对聚谷氨酸衍生物的催化速率的差异效应。叶酸的4-氨基取代增加了单谷氨酸衍生物的结合速率，但严重损害了二谷氨酸衍生物的催化作用。蝶酰鸟氨酸衍生物是第一个被鉴定的有效的和特异的folylpolyglutamate合成酶的底物，并且可以作为反应中间体的类似物。具有取代肽键的四面体化学的其他叶酸衍生物，如蝶酰基-7-谷氨酰基-[N，CH 2-NH]-谷氨酸，是比鸟氨酸衍生物有效的抑制剂。
Revised Manuscript Received September 25, 1986 abstract: A variety of folate analogues were synthesized to explore the specificity of the folate binding site of hog liver folylpolyglutamate synthetase and the requirements for catalysis. Modifications of the internal and terminal glutamate moieties of folate cause large drops in on rates and/or affinity for the protein. The only exceptions are glutamine, homocysteate, and ornithine analogues, indicating a less stringent specificity around the-carbonof glutamate. It is proposed that initial folate bindingto the enzyme involves low-affinity interactions at a pterin and a glutamate site and that the first glutamate bound is the internal residue adjacent to the benzoyl group. Processive movement of the polyglutamate chain through the glutamate site and a possible conformational change in the protein when the terminal residue is bound would result in tight binding and would position the-carboxyl of the terminal glutamate inthe correct position for catalysis. Steric limitations imposed on the internal glutamate residues that loop out and additional steric constraints imposed by binding of different pterin moieties would be expected to effect slight conformational changes in the protein and/or the terminal glutamate and would explain the decrease in on rate and catalytic rate with increased polyglutamate chain length, and the differential effect of one-carbon substitution on the catalytic rate with polyglutamate derivatives. The 4-amino substitution of folate increases the on rate for monoglutamate derivatives but severely impairs catalysis with diglutamate derivatives. Pteroylornithine derivatives are the first potent and specific inhibitorsof folylpolyglutamate synthetase to be identified and may act as analogues of reaction intermediates. Other folate derivatives with tetrahedral chemistry replacing the peptide bond, such as pteroyl-7-glutamyl-[i/', CH2-NH]-glutamate, effective inhibitors than the ornithine derivatives.