Jefcoate C R
J Biol Chem. 1975 Jun 25;250(12):4663-70.
Adrenal mitochondrial cytochrome P-450 which functions in cholesterol side chain cleavage (P-450scc) exhibited type I (lambdamax 385, lambdamin 420 nm) and inverse type I (lambdamin 385, lambdamax 420 nm) difference spectra with several steroids. The magnitude and type of response were dependent on the particular steroid and on the extent to which cholesterol was bound to the cytochrome in the intact mitochondrion. the inverse type I difference spectrum induced by 3beta-hydroxy-pregn-5-ene-20-one (pregnenolone) was dependent on the proportion of high spin cholesterol-cytochrome P-450scc complexes. With rat adrenal mitochondria cholest-5-ene-3beta, 20alpha-diol (20alpha-hydroxycholesterol) invariably induced a smaller inverse type I response and, under conditions where cytochrome P-450scc was nearly free of cholesterol, even produced a small type I response. Two distinct steroid binding sites on cytochrome P-450scc were detected by, respectively, the slow type I response to cholest-5-ene-3beta, 25-diol (25-hydroxycholesterol) and the rapid type I response to a subsequent addition of cholest-5-ene-3beta, 20alpha, 22 R-triol (20alpha, 22R-dihydroxycholesterol). The relative proportions of the spectral responses to these steroids were dependent on the previous extent of adrenal activation by adrenocorticotropic hormone (ACTH), because this stimulatory process altered the combination of mitochondrial cholesterol with cytochrome P-450scc. It is proposed that the two steroid binding sites on cytochrome P-450scc interact with steroids in the following way: site I binds cholesterol, 25-hydroxycholesterol, and 20alpha, 22R-dihydroxycholesterol with formation of a partially high spin cytochrome; site II binds both pregnenolone and 20alpha-OH cholesterol resulting in a low spin cytochrome. Interactions between sites I and II are not competitive, and occupancy of site II ensures a low spin state irrespective of the occupancy of site I. A second mode of interaction by 20alpha, 22R-dihydroxycholesterol stabilizes a high spin cytochrome and is competitive with site II binding by 20alpha-hydroxycholesterol or pregnenolone. Formation of a maximally high spin cytochrome follows occupancy by 20alpha, 22R-dihydroxycholesterol at both sites.
肾上腺线粒体中参与胆固醇侧链裂解的细胞色素P - 450（P - 450scc）与几种甾体呈现出I型（最大吸收波长385nm，最小吸收波长420nm）和反向I型（最小吸收波长385nm，最大吸收波长420nm）差异光谱。响应的大小和类型取决于特定的甾体以及胆固醇在完整线粒体中与细胞色素结合的程度。由3β - 羟基 - 孕 - 5 - 烯 - 20 - 酮（孕烯醇酮）诱导的反向I型差异光谱取决于高自旋胆固醇 - 细胞色素P - 450scc复合物的比例。对于大鼠肾上腺线粒体，胆甾 - 5 - 烯 - 3β,20α - 二醇（20α - 羟基胆固醇）总是诱导出较小的反向I型响应，并且在细胞色素P - 450scc几乎不含胆固醇的条件下，甚至产生小的I型响应。分别通过对胆甾 - 5 - 烯 - 3β,25 - 二醇（25 - 羟基胆固醇）的缓慢I型响应和对随后添加的胆甾 - 5 - 烯 - 3β,20α,22R - 三醇（20α,22R - 二羟基胆固醇）的快速I型响应，检测到细胞色素P - 450scc上两个不同的甾体结合位点。对这些甾体的光谱响应的相对比例取决于先前促肾上腺皮质激素（ACTH）对肾上腺的激活程度，因为这种刺激过程改变了线粒体胆固醇与细胞色素P - 450scc的结合情况。有人提出，细胞色素P - 450scc上的两个甾体结合位点与甾体的相互作用方式如下：位点I结合胆固醇、25 - 羟基胆固醇和20α,22R - 二羟基胆固醇，形成部分高自旋细胞色素；位点II结合孕烯醇酮和20α - 羟基胆固醇，导致低自旋细胞色素。位点I和位点II之间的相互作用不是竞争性的，并且位点II的占据确保了低自旋状态，而与位点I的占据情况无关。20α,22R - 二羟基胆固醇的第二种相互作用模式稳定了高自旋细胞色素，并且与20α - 羟基胆固醇或孕烯醇酮在位点II的结合具有竞争性。两个位点都被20α,22R - 二羟基胆固醇占据后，形成最大程度的高自旋细胞色素。
