The highly stereoselective oxidation of polyunsaturated fatty acids by cytochrome P450BM-3
The highly stereoselective oxidation of polyunsaturated fatty acids by cytochrome P450BM-3
复制标题
DOI：
10.1074/jbc.271.37.22663
复制
发表时间：
1996-09-13
影响因子：
4.8
通讯作者：
Peterson, JA
中科院分区：
文献类型：
--
作者：
Capdevila, JH;Wei, SZ;Peterson, JA
细胞色素P450 BM-3催化花生四烯酸的NADPH依赖性代谢为几乎对映体纯的18（R）-羟基二十碳四烯酸和14（S），15（R）-环氧二十碳三烯酸（分别占总产物的80%和20%），P450 BM-3在30 ℃下以3.2 +/-0.4 μ mol/min/nmol的速率氧化花生四烯酸，这是有史以来报道的最快的NADPH依赖性P450催化反应，脂肪酸、氧和NADPH以约1：1：1的摩尔比利用，表明电子传递与单加氧有效偶联。二十碳五烯酸和二十碳三烯酸是两种花生四烯酸类似物，其C-15-C-18碳的性质不同，也被P450 BM-3积极代谢（30 ° C时分别为1.4 +/- 0.2和2.9 +/- 0.1 μ mol/min/nmol）。而二十碳五烯酸的17，18-烯键以几乎绝对的区域和立体化学选择性被环氧化成17（S），18（R）-环氧二十碳四烯酸（大于或等于总产物的99%，97%光学纯度），P450 BM-3在二十碳三烯酸ω-1、ω-2和ω-3sp（3）碳的羟基化过程中仅具有中等的区域选择性，其中17-、18-和19-羟基二十碳三烯酸分别以2.4：2.2：1的比例形成。基于上述和花生四烯酸结合的P450 BM-3的模型，我们提出：1）通过P450 BM-3形成能够烯键环氧化和sp（3）碳羟基化的单一氧化剂物质，和2）产物化学，因此，催化结果关键地取决于负责底物结合的活性位点空间坐标和血红素结合的活性氧与受体碳键之间的有效取向。
Cytochrome P450BM-3 catalyzes NADPH-dependent metabolism of arachidonic acid to nearly enantiomerically pure 18(R-hydroxyeicosatetraenoic acid and 14(S),15(R)-epoxyeicosatrienoic acid (80 and 20% of total products, respectively), P450BM-3 oxidizes arachidonic acid with a rate of 3.2 +/- 0.4 mu mol/min/nmol at 30 degrees C, the fastest ever reported for an NADPH dependent, P450-catalyzed reaction, Fatty acid, oxygen, and NADPH are utilized in an approximately 1:1:1 molar ratio, demonstrating efficient coupling of electron transport to monooxygenation.Eicosapentaenoic and eicosatrienoic acids, two arachidonic acid analogs that differ in the properties of the C-15-C-18 carbons, are also actively metabolized by P450BM-3 (1.4 +/- 0.2 and 2.9 +/- 0.1 mu mol/min/nmol at 30 degrees C, respectively). While the 17,18-olefinic bond of eicosapentaenoic acid is epoxidized with nearly absolute regio- and stereochemical selectivity to 17(S),18(R)-epoxyeicosatetraenoic acid (greater than or equal to 99% of total products, 97% optical purity), P450BM-3 is only moderately regioselective during hydroxylation of the eicosatrienoic acid omega-1, omega-2, and omega-3 sp(3) carbons, with 17-, 18-, and 19-hydroxyeicosatrienoic acid formed in a ratio of 2.4:2.2:1, respectively.Based on the above and on a model of arachidonic acid-bound P450BM-3, we propose: 1) the formation by P450BM-3 of a single oxidant species capable of olefinic bond epoxidation and sp(3) carbon hydroxylation and 2) that product chemistry and, thus, catalytic outcome are critically dependent on active site spatial coordinates responsible for substrate binding and productive orientation between heme-bound active oxygen and acceptor carbon bond(s).