蛋白质胰蛋白酶水解过程双位点漏切肽段的质谱鉴定  

Identification of Tryptic Peptides with Double Missed Cleavage Sites by Mass Spectrometry

作　　者：王勇[1] 李水明[1] 何曼文[2] 邹永东[2] 

机构地区：[1]深圳大学生命科学学院,深圳市海洋生物资源与生态环境重点实验室,广东深圳518060 [2]深圳大学生命科学学院,深圳市微生物基因工程重点实验室,广东深圳518060

出　　处：《质谱学报》2013年第1期29-34,共6页Journal of Chinese Mass Spectrometry Society

基　　金：国家自然科学基金(31070731);深圳市科技研发资金科技条件平台计划重点实验室项目(CXB201005240008A)资助

摘　　要：为说明胰蛋白酶漏切位点数目设置对蛋白质搜库结果的影响,采用基质辅助激光解吸电离-串联飞行时间质谱(MALDI-TOF/TOF)高置信度地鉴定了细胞色素C和大豆脱落胁迫成熟蛋白。在细胞色素C和大豆脱落胁迫成熟蛋白的胰酶水解液中分别发现了2个和4个双胰酶漏切位点肽段,确认了文献中报道的当赖氨酸和赖氨酸相邻或赖氨酸、精氨酸各自与天冬氨酸或者谷氨酸相邻时,会引起胰蛋白酶的漏切。此外,还发现组氨酸-赖氨酸-异亮氨酸/丙氨酸结构也有此现象。本研究中检测出漏切肽段并没有明显提高序列覆盖率,但显著增加了蛋白质鉴定结果的确定性。在实际样品分析中如将漏切数设置为1有可能产生肽段鉴定的假阴性。To investigate the influence of missed cleavage numbers of trypsin for database searching results, Cytochrome C and abscisic stress ripening-like protein were identified by MALDI-TOF/TOF mass spectrometry. Two and four tryptic peptides with double missed cleavage were found in the digests of cytochrome C and of abscisic stress ripening-like protein, respectively. It was found that two missed cleavage will happe when two lysine residues neighbor to each other or lysines and arginines neighbor with an aspartie acid or glutamic acid residue. It was observed that this phenomenon was related to histine-lysine-isoleunine/alanine sequence. In this study, the certainty of protein identification rather than protein coverage was significantly improved when tryptic peptides with double missed clearage sites were identified, which indicate that false negative of peptide identification will hap pen when the number of missed cleavage site was set to 1.

关 键 词：胰蛋白酶 基质辅助激光解吸电离-串联飞行时间质谱 细胞色素C 大豆脱落胁迫成熟蛋白 

分 类 号：O657.63[理学—分析化学]