Stereochemistry of the chloroperoxidase active site: crystallographic and molecular-modeling studies
Stereochemistry of the chloroperoxidase active site: crystallographic and molecular-modeling studies
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DOI：
10.1016/s1074-5521(98)90003-5
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发表时间：
1998-09-01
影响因子：
--
通讯作者：
Poulos, TL
中科院分区：
文献类型：
--
作者：
Sundaramoorthy, M;Terner, J;Poulos, TL
背景：氯过氧化物酶(CPO)是已知的用途最广的血红素酶。它催化活化的C-H键的氯化，以及过氧化物酶、过氧化氢酶和细胞色素P450的反应，包括对映选择性环氧化。CPO含有近端的亚铁血红素硫酸盐配体，如P450，以及远端的极性口袋，如过氧化物酶。底物结合位置是由血红素上方的开口形成的，该开口使有机底物能够接近活化的氧铁基氧原子。与其他过氧化物酶不同的是，CPO使用谷氨酸酸碱催化剂，而不是组氨酸残基。结果：确定了CPO与外源配体一氧化碳、一氧化氮、氰化物和硫氰酸盐的络合物的晶体结构。远端口袋根据大小和pK(A)区分配体。精炼的CPO配体结构表明，具有固定的谷氨酸酸碱催化剂的刚性活性中心结构。以顺-β-甲基苯乙烯为底物的环氧化反应的分子模拟和动力学模拟为理解环氧化反应的对映选择性提供了结构和能量基础。结论：不同的GPO配体结构为以Glu183为酸碱催化剂的中间体I的形成提供了详细的立体化学机理的基础。观察到的活性中心的刚性也解释了CPO化合物I的相对不稳定性和HOCl氯化物种的形成。CPO的能量学-底物/产物分子模拟为CPO的P450型对映体选择性环氧化活性提供了理论依据。
Background: Chloroperoxidase (CPO) is the most versatile of the known heme enzymes. It catalyzes chlorination of activated C-H bonds, as well as peroxidase, catalase and cytochrome P450 reactions, including enantioselective epoxidation. CPO contains a proximal heme-thiolate ligand, like P450, and polar distal pocket, like peroxidase. The substrate-binding site is formed by an opening above the heme that enables organic substrates to approach the activated oxoferryl oxygen atom. CPO, unlike other peroxidases, utilizes a glutamate acid-base catalyst, rather than a histidine residue.Results: The crystal structures of CPO complexed with exogenous ligands, carbon monoxide, nitric oxide, cyanide and thiocyanate, have been determined. The distal pocket discriminates ligands on the basis of size and pK(a). The refined CPO-ligand structures indicate a rigid active-site architecture with an immobile glutamate acid-base catalyst. Molecular modeling and dynamics simulations of CPO with the substrate cis-beta-methylstyrene and the corresponding epoxide products provide a structural and energetic basis for understanding the enantioselectivity of CPO-catalyzed epoxidation reactions.Conclusions: The various GPO-ligand structures provide the basis for a detailed stereochemical mechanism of the formation of the intermediate compound I, in which Glu183 acts as an acid-base catalyst. The observed rigidity in the active site also explains the relative instability of CPO compound I and the formation of the HOCl chlorinating species. Energetics of CPO-substrate/product molecular modeling provides a theoretical basis for the P450-type enantioselective epoxidation activities of CPO.