2DE-MS中膜蛋白质组的研究进展

2-维凝胶电泳(2DE)具有高分辨率、高通量等特点,已被广泛地用于蛋白质组的研究.然而,2DE-MS在膜蛋白质组学研究方面却有其局限性,主要因为:膜蛋白具有低丰度、难溶、等电点时易沉淀、难酶解等特点.然而随着亚细胞分离技术和直接的生化方法富集等技术的发展,低丰度问题得到了极大的改善;增溶剂(尿素,硫脲),新的两性离子和非离子去垢剂,以及有机溶剂等的利用极大地改善了膜蛋白质组的溶解性能;同时,一些新的2DE技术的利用扩大了常规2DE的分离范围.在膜蛋白裂解方面,将酶解法与化学法(CNBr)相结合,另外先进的质谱技术的发展使得膜蛋白质组的研究在最近几年取得了较大的发展.现对2DE-MS途径中,膜的富集、膜蛋白的提取、分离、酶解、鉴定方面的进展进行综述.

The Application of 2DE-MS to Membrane Proteomics

DE has the property of high resolution, high throughout , so it has been used for the study of proteome. However, it has great limitation in the study of membrane proteome, mainly because of three reasons: low copy, poor solubility, easy precipitation (at their isoelectric point), difficult digestion of membrane proteins. However, with the development of subcellular fractionation technology and directed biochemical enrichments technology, which have enriched the membrane proteins, the use of chaotrope (Urea, Thiourea), new zwitterionic detergent, non ion detergent, and some organic solvents, which has improved the solution of membrane proteins, at the same time, in addition to IPG IEF /SDS PAGE, some new methods (such as cation/anion electrophoresis, two dimensional blue native electrophoresis), which have extended the separation of membrane proteins, furthermore , the combination of proteases and chemical cleavage methods, which has improved the sequence coverage of the hydrophobic membrane domain, at the same time, Mass Spectrometry with the high sensitive, high resolution which can be used to identify the membrane proteins accurately, so the study of membrane proteomics has made great improvement. The enrichment, extraction, separation, digestion, identification of membrane proteins in the 2DE/MS were introduced.