Nisar S, Lane C S, Wilderspin A F, Welham K J, Griffiths W J, Patterson L H
Department of Pharmaceutical & Biological Chemistry, The School of Pharmacy, University of London, 29-39 Brunswick Square, London, United Kingdom.
Drug Metab Dispos. 2004 Apr;32(4):382-6. doi: 10.1124/dmd.32.4.382.
Nanoscale reversed-phase liquid chromatography (LC) combined with electrospray ionization-tandem mass spectrometry (ESI-MS/MS) has been used as a method for the direct identification of multiple cytochrome P450 (P450) isoforms found in male and female rat liver. In this targeted proteomic approach, rat liver microsomes were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by in-gel tryptic digestion of the proteins present in the 48- to 62-kDa bands. The resultant peptides were extracted and analyzed by LC-ESI-MS/MS. P450 identifications were made by searching the MS/MS data against a rat protein database containing 21,576 entries including 47 P450s using Sequest software (Thermo Electron, Hemel Hempstead, UK). Twenty-four P450 isoforms from the subfamilies 1A, 2A, 2B, 2C, 2D, 2E, 3A, 4A, 4F, CYP17, and CYP19 were positively identified in rat liver.
纳米级反相液相色谱（LC）与电喷雾电离串联质谱（ESI-MS/MS）联用，已被用作直接鉴定雄性和雌性大鼠肝脏中多种细胞色素P450（P450）同工型的方法。在这种靶向蛋白质组学方法中，大鼠肝脏微粒体先进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳，然后对48至62 kDa条带中的蛋白质进行胶内胰蛋白酶消化。提取所得肽段并通过LC-ESI-MS/MS进行分析。使用Sequest软件（英国赫默尔亨普斯特德的热电电子公司），通过将MS/MS数据与包含21576个条目（其中包括47种P450）的大鼠蛋白质数据库进行比对来鉴定P450。在大鼠肝脏中阳性鉴定出了来自1A、2A、2B、2C、2D、2E、3A、4A、4F、CYP17和CYP19亚家族的24种P450同工型。
