The F-G loop region of cytochrome P450scc (CYP11A1) interacts with the phospholipid membrane
The F-G loop region of cytochrome P450scc (CYP11A1) interacts with the phospholipid membrane
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DOI：
10.1016/j.bbamem.2003.09.007
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发表时间：
2003-10-31
影响因子：
3.4
通讯作者：
Tuckey, RC
中科院分区：
文献类型：
--
作者：
Headlam, MJ;Wilce, MCJ;Tuckey, RC
细胞色素P450 SCC（CYP 11 A1）是一种附着在线粒体内膜内表面的蛋白质，它使用膜相的胆固醇作为类固醇激素合成的第一步底物。我们研究了CYP 11 A1与膜相互作用的机制。CYP 11 A1和其他两种线粒体细胞色素P450的疏水性特征，加上以CYP 2C 5为模板的CYP 11 A1的模型结构，表明CYP 11 A1与膜具有单调的结合，其可能涉及A'螺旋和F-G环。A'螺旋的缺失降低了与细菌膜组分结合的表达的CYP 11 A1的比例，表明A'螺旋在膜结合中的作用。然而，在该螺旋的位置24（L24 C）处引入半胱氨酸残基并随后用荧光探针N '-（7-硝基苯-2-草酸，3-二唑-4-基）乙二胺（NBD）标记未能显示膜定位。半胱氨酸诱变剂和荧光标记的其他残基上出现的远端表面的CYP 11 A1模型显示，V212 C和L219 C具有增强的荧光和蓝移后的突变体CYP 11 A1与磷脂囊泡的协会。这暗示了这些位于F-G环中的残基在膜结合后变得定位于更疏水的环境。丙烯酰胺对CYP 11 A1中色氨酸残基的淬灭分析表明，至少有一个，可能有两个色氨酸残基参与膜结合。我们的结论是，CYP 11 A1与膜介导的，至少部分，由F-G环区的单调关联。(C)2003 Elsevier B. V.保留所有权利。
Cytochrome P450scc (CYP11A1) is a protein attached to the inner surface of the inner mitochondrial membrane that uses cholesterol from the membrane phase as its substrate for the first step in steroid hormone synthesis. We investigated the mechanism by which CYP11A1 interacts with the membrane. Hydrophobicity profiles of CYP11A1 and two other rnitochondrial cytochromes P450, plus a model structure of CYP11A1 using CYP2C5 as template, suggest that CYP11A1 has a monotopic association with the membrane which may involve the A' helix and the F-G loop, Deletion of the A' helix reduced the proportion of expressed CYP11A1 associated with the bacterial membrane fraction, indicating a role for the A' helix in membrane binding. However, introduction of a cysteine residue in this helix at position 24 (L24C) and subsequent labelling with the fluorescent probe N'-(7-nitrobenz-2-oxal,3-diazol-4-yl)ethylenediamine (NBD) failed to show a membrane localisation. Cysteine mutagenests and fluorescent labelling of other residues appearing on the distal surface of the CYP11A1 model revealed that V212C and L219C have enhanced fluorescence and a blue shift following association of the mutant CYP11A1 with phospholipid vesicles. This inchcates that these residues, which are located in the F-G loop, become localised to a more hydrophobic environment following membrane binding. Analysis of the quenching of tryptophan residues in CYP11A1 by acrylamide indicates that at least one and probably two tryptophans are involved in membrane binding. We conclude that CYP11A1 has a monotopic association with the membrane that is mediated, at least in part, by the F-G loop region. (C) 2003 Elsevier B.V. All rights reserved.