Structural Basis for Three-step Sequential Catalysis by the Cholesterol Side Chain Cleavage Enzyme CYP11A1
Structural Basis for Three-step Sequential Catalysis by the Cholesterol Side Chain Cleavage Enzyme CYP11A1
复制标题
DOI：
10.1074/jbc.m110.188433
复制
发表时间：
2011-02-18
影响因子：
4.8
通讯作者：
Pikuleva, Irina A.
中科院分区：
文献类型：
--
作者：
Mast, Natalia;Annalora, Andrew J.;Pikuleva, Irina A.
线粒体细胞色素P450 11 A1（CYP 11 A1或P450 11 A1）是唯一已知的酶，它可以切割胆固醇的侧链，产生所有类固醇激素的前体-烯醇酮。孕烯醇酮是通过三个连续的单氧化反应形成的，这些反应涉及22 R-羟基胆固醇（22 HC）和20 α，22 R-二羟基胆固醇的逐步产生，然后是C20-C22键的裂解。在这里，我们提出了2.5埃晶体结构的CYP 11 A1与第一个反应中间体，22 HC的复合物。CYP 11 A1中的活性位点空腔代表了一个从蛋白质表面延伸到血红素基团（催化位点）的长弯管。22 HC占据了三分之二的空腔，其中22 R-羟基最接近血红素，距离铁2.56埃。活性中心入口处的空间没有被22 HC占据，而是充满了有序的水分子。由这些水分子形成的网络允许22 HC 3 β-羟基的“软”识别。这种模式的22 HC结合表明穿梭的甾醇中间体之间的活性位点入口和血红素基团在三步反应。22 HC的平移自由和扭转运动的脂肪族尾巴的解决方案的研究支持。CYP 11 A1 - 22 HC共复合物还提供了对酶的严格底物特异性和高催化效率的结构基础的深入了解，并突出了线粒体P450参与氧化还原伴侣相互作用的保守结构基序。
Mitochondrial cytochrome P450 11A1 (CYP11A1 or P450 11A1) is the only known enzyme that cleaves the side chain of cholesterol, yielding pregnenolone, the precursor of all steroid hormones. Pregnenolone is formed via three sequential monooxygenation reactions that involve the progressive production of 22R-hydroxycholesterol (22HC) and 20 alpha,22R-dihydroxycholesterol, followed by the cleavage of the C20-C22 bond. Herein, we present the 2.5-angstrom crystal structure of CYP11A1 in complex with the first reaction intermediate, 22HC. The active site cavity in CYP11A1 represents a long curved tube that extends from the protein surface to the heme group, the site of catalysis. 22HC occupies two-thirds of the cavity with the 22R-hydroxyl group nearest the heme, 2.56 angstrom from the iron. The space at the entrance to the active site is not taken up by 22HC but filled with ordered water molecules. The network formed by these water molecules allows the "soft" recognition of the 22HC 3 beta-hydroxyl. Such a mode of 22HC binding suggests shuttling of the sterol intermediates between the active site entrance and the heme group during the three-step reaction. Translational freedom of 22HC and torsional motion of its aliphatic tail are supported by solution studies. The CYP11A1-22HC co-complex also provides insight into the structural basis of the strict substrate specificity and high catalytic efficiency of the enzyme and highlights conserved structural motifs involved in redox partner interactions by mitochondrial P450s.