Goyal Sandeep, Xiao Yi, Porter Ned A, Xu Libin, Guengerich F Peter
Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, TN 37232.
Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235.
J Lipid Res. 2014 Sep;55(9):1933-43. doi: 10.1194/jlr.M051508. Epub 2014 Jul 13.
Cytochrome P450 (P450 or CYP) 46A1 is expressed in brain and has been characterized by its ability to oxidize cholesterol to 24S-hydroxycholesterol. In addition, the same enzyme is known to further oxidize 24S-hydroxycholesterol to the 24,25- and 24,27-dihydroxy products, as well as to catalyze side-chain oxidations of 7α-hydroxycholesterol and cholestanol. As precursors in the biosynthesis of cholesterol, 7-dehydrocholesterol has not been found to be a substrate of P450 46A1 and desmosterol has not been previously tested. However, 24-hydroxy-7-dehydrocholesterol was recently identified in brain tissues, which prompted us to reexamine this enzyme and its potential substrates. Here we report that P450 46A1 oxidizes 7-dehydrocholesterol to 24-hydroxy-7-dehydrocholesterol and 25-hydroxy-7-dehydrocholesterol, as confirmed by LC-MS and GC-MS. Overall, the catalytic rates of formation increased in the order of 24-hydroxy-7-dehydrocholesterol < 24-hydroxycholesterol < 25-hydroxy-7-dehydrocholesterol from their respective precursors, with a ratio of 1:2.5:5. In the case of desmosterol, epoxidation to 24S,25-epoxycholesterol and 27-hydroxylation was observed, at roughly equal rates. The formation of these oxysterols in the brain may be of relevance in Smith-Lemli-Opitz syndrome, desmosterolosis, and other relevant diseases, as well as in signal transduction by lipids.
细胞色素P450（P450或CYP）46A1在大脑中表达，其特征在于能够将胆固醇氧化为24S-羟基胆固醇。此外，已知同一种酶能将24S-羟基胆固醇进一步氧化为24,25-二羟基产物和24,27-二羟基产物，还能催化7α-羟基胆固醇和胆甾烷醇的侧链氧化。作为胆固醇生物合成的前体，尚未发现7-脱氢胆固醇是P450 46A1的底物，而羊毛甾醇此前也未经过测试。然而，最近在脑组织中发现了24-羟基-7-脱氢胆固醇，这促使我们重新审视这种酶及其潜在底物。在此我们报告，经液相色谱-质谱联用（LC-MS）和气相色谱-质谱联用（GC-MS）证实，P450 46A1将7-脱氢胆固醇氧化为24-羟基-7-脱氢胆固醇和25-羟基-7-脱氢胆固醇。总体而言，从各自的前体生成这些产物的催化速率按24-羟基-7-脱氢胆固醇＜24-羟基胆固醇＜25-羟基-7-脱氢胆固醇的顺序增加，比例为1:2.5:5。对于羊毛甾醇，观察到其以大致相等的速率环氧化为24S,25-环氧胆固醇并发生27-羟基化。这些氧甾醇在大脑中的形成可能与史密斯-利姆利-奥皮茨综合征、羊毛甾醇病及其他相关疾病有关，也与脂质的信号转导有关。
