Anal Chem. 2010 Dec 1;82(23):9622-5. doi: 10.1021/ac1023099. Epub 2010 Nov 3.
In this study, a facile system for membrane proteome profiling was established, in which membrane proteins were solubilized by formic acid, online digested by a pepsin-based immobilized enzyme reactor (pepsin-IMER), and analyzed by strong cation exchange and microflow reversed-phase liquid chromatography with electrospray ionization tandem mass spectrometry (SCX-μRPLC-ESI-MS/MS). Under optimized conditions, such a system showed excellent compatibility between all crucial steps and was successfully applied for analyzing integral membrane proteins extracted from rat liver microsomes. Out of the 235 unique proteins positively identified, 39% (91/235) were annotated as membrane proteins with one or more transmembrane domains (TMDs). It is anticipated that the efficient sample treatment and the relevant online analytical system might provide a promising tool for automated and comprehensive profiling of membrane proteomes.
在本研究中，建立了一种用于膜蛋白质组学分析的简易系统，其中膜蛋白通过甲酸溶解，在线用基于胃蛋白酶的固定化酶反应器（胃蛋白酶 IMER）进行消化，然后通过强阳离子交换和微流反向相液相色谱与电喷雾串联质谱（SCX-μRPLC-ESI-MS/MS）进行分析。在优化条件下，该系统在所有关键步骤之间具有出色的兼容性，并成功应用于分析从大鼠肝微粒体中提取的完整膜蛋白。在鉴定的 235 个独特蛋白质中，有 39%（91/235）被注释为具有一个或多个跨膜结构域（TMDs）的膜蛋白。预计这种有效的样品处理和相关的在线分析系统可能为自动化和全面的膜蛋白质组学分析提供一种有前途的工具。
