Development and evaluation of an entirely solution-based combinative sample preparation method for membrane proteomics
Development and evaluation of an entirely solution-based combinative sample preparation method for membrane proteomics
复制标题
膜蛋白质组学完全基于溶液的组合样品制备方法的开发和评估
DOI：
10.1016/j.ab.2012.09.023
复制
发表时间：
2013-01-01
影响因子：
2.9
通讯作者：
Liang, Songping
中科院分区：
文献类型：
--
作者：
Lin, Yong;Liu, Hui;Liang, Songping
许多膜蛋白，特别是整体膜蛋白的疏水性给其分析带来了很大的困难。为了提高膜蛋白的分析水平，本研究提出了一种基于全溶液的组合样品制备（CSP）方法，并对其在大鼠肝膜蛋白质组学鸟枪法分析中的应用进行了评价。该CSP方法综合利用了十二烷基硫酸钠（SDS）裂解膜和溶解疏水膜蛋白的强能力，优化后的丙酮沉淀法在样品清理和蛋白质回收方面的高效率，以及脱氧胆酸钠（SDC）在提高蛋白质溶解/消化以及与胰蛋白酶活性相容方面的优势。与其他两种具有代表性的样品制备方法sdc辅助溶膜法和管凝胶法相比，新建立的CSP方法在回收和鉴定疏水肽、较大肽和具有多个跨膜结构域的高度疏水膜蛋白方面具有优势。CSP方法具有操作简单、成本低、适用于处理各种体积的蛋白质样品，特别是大体积的蛋白质样品等特点，在膜蛋白质组的质谱分析中具有潜力。(C) 2012爱思唯尔公司版权所有。
The hydrophobic nature of many membrane proteins, especially integral membrane proteins, brings great difficulties to their analysis. To improve the analysis of membrane proteins, an entirely solution-based combinative sample preparation (CSP) method was developed and its application to the shotgun analysis of rat liver membrane proteomes was evaluated in this study. This CSP method comprehensively uses the strong ability of sodium dodecyl sulfate (SDS) to lyse the membranes and solubilize hydrophobic membrane proteins, the high efficiency of the optimized acetone precipitation method in sample cleanup and protein recovery, and the advantages of sodium deoxycholate (SDC) in improving protein solubilization/digestion as well as being compatible with trypsin activity. Compared with two other representative sample preparation methods, the SDC-assisted membrane-lysing method and the tube gel method, the newly established CSP method exhibited superiority in the recovery and identification of hydrophobic peptides, larger peptides, and highly hydrophobic membrane proteins with multiple transmembrane domains. The CSP method has characteristics of easy operation, low cost, and suitability for treating protein samples in various volumes, particularly large volumes, thereby having potential in the analysis of membrane proteomes with mass spectrometry. (C) 2012 Elsevier Inc. All rights reserved.