Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis

	Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis
	Liu, Fangjie 1,2; Ye, Mingliang 1; Pan, Yanbo 1,2; Zhang, Yi 1,2; Bian, Yangyang 1,2; Sun, Zhen 1,2; Zhu, Jun 1,2; Cheng, Kai 1,2; Zou, Hanfa 1
刊名	analytical chemistry
	2014-07-15
卷号	86 期号:14 页码:6786-6791
ISSN号	0003-2700
通讯作者	邹汉法 ; 叶明亮
产权排序	待补充
合作状况	英
英文摘要	conventional sample preparation protocols for phosphoproteome analysis require multiple time-consuming and labor-intensive steps, including cell lysis, protein extraction, protein digestion, and phosphopeptide enrichment. in this study, we found that the presence of a large amount of trypsin in the sample did not interfere with phosphopeptide enrichment and subsequent lc-ms/ms analysis. taking advantage of fast digestion achieved with high trypsin-to-protein ratio, we developed a novel concurrent lysis-digestion method for phosphoproteome analysis. in this method, the harvested cells were first placed in a lysis buffer containing a huge amount of trypsin. after ultrasonication, the cells were lysed and the proteins were efficiently digested into peptides within one step. thereafter, tryptic digest was subjected to phosphopeptide enrichment, in which unphosphorylated peptides, trypsin, and other components incompatible with lc-ms/ms analysis were removed. compared with conventional methods, better phosphoproteome coverage was achieved in this new one-step method. because protein solubilization and cell lysis were facilitated by fast protein digestion, the complete transformation of cell pellets into the peptide mixture could be finished within 25 min, while it would take at least 16 h for conventional methods. hence, our method, which integrated cell lysis, protein extraction, and protein digestion into one step, is rapid and convenient. it is expected to have broad applications in phosphoproteomics analysis.
学科主题	物理化学
WOS标题词	science & technology ; physical sciences
类目[WOS]	chemistry, analytical
研究领域[WOS]	chemistry
关键词[WOS]	immobilized trypsin digestion ; intensity focused ultrasound ; proteomics ; chromatography ; impact ; mass
收录类别	SCI
资助信息	1,1
原文出处	6791
语种	英语
WOS记录号	WOS:000339227400010
公开日期	2014-09-11
内容类型	期刊论文
源URL	[http://159.226.238.44/handle/321008/119725]
专题	大连化学物理研究所_中国科学院大连化学物理研究所
作者单位	1.Chinese Acad Sci, Dalian Inst Chem Phys, Key Lab Separat Sci Analyt Chem, Dalian 116023, Liaoning, Peoples R China 2.Chinese Acad Sci, Grad Sch, Beijing 100049, Peoples R China
推荐引用方式 GB/T 7714	Liu, Fangjie,Ye, Mingliang,Pan, Yanbo,et al. Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis[J]. analytical chemistry,2014,86(14):6786-6791.
APA	Liu, Fangjie.,Ye, Mingliang.,Pan, Yanbo.,Zhang, Yi.,Bian, Yangyang.,...&Zou, Hanfa.(2014).Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis.analytical chemistry,86(14),6786-6791.
MLA	Liu, Fangjie,et al."Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis".analytical chemistry 86.14(2014):6786-6791.