Wang Sheng, Wei Bangguo, Yang Pengyuan, Chen Gang
School of Pharmacy, Institute of Biomedical Sciences, Fudan University, Shanghai, PR China.
Proteomics. 2008 Nov;8(22):4637-41. doi: 10.1002/pmic.200800401.
In this report, alternating current-assisted on-plate proteolysis has been developed for rapid peptide mapping. Protein solutions containing trypsin were allowed to digest directly on the spots of a stainless steel MALDI plate with the assistance of low-voltage alternating current electricity. Alternating current (AC) was allowed to pass through the protein solutions via the MALDI plate and a platinum disc electrode. The feasibility and performance of the novel proteolysis approach were investigated by the digestion of BSA and cytochrome c (Cyt-c). It was demonstrated that AC substantially enhanced the efficiency of proteolysis and the digestion time was significantly reduced to 5 min. The digests were identified by MALDI-TOF MS with sequence coverages of 42% (BSA) and 77% (Cyt-c) that were comparable to those obtained by using conventional in-solution tryptic digestion. The present proteolysis strategy is simple and efficient, offering great promise for MALDI-TOF MS peptide mapping.
在本报告中，已开发出交流电辅助板上蛋白水解技术用于快速肽图谱分析。含有胰蛋白酶的蛋白质溶液在低压交流电的辅助下，直接在不锈钢基质辅助激光解吸电离（MALDI）板的斑点上进行消化。交流电通过MALDI板和铂盘电极穿过蛋白质溶液。通过对牛血清白蛋白（BSA）和细胞色素c（Cyt-c）的消化研究了这种新型蛋白水解方法的可行性和性能。结果表明，交流电显著提高了蛋白水解效率，消化时间显著缩短至5分钟。通过基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）对消化产物进行鉴定，序列覆盖率分别为42%（BSA）和77%（Cyt-c），与使用传统溶液内胰蛋白酶消化获得的覆盖率相当。目前的蛋白水解策略简单高效，为MALDI-TOF MS肽图谱分析提供了广阔前景。
