Resin-assisted enrichment of N-terminal peptides for characterizing proteolytic processing.
Resin-assisted enrichment of N-terminal peptides for characterizing proteolytic processing.
复制标题
DOI：
10.1021/ac401000q
复制
发表时间：
2013-07-16
影响因子：
7.4
通讯作者：
Qian, Wei-Jun
中科院分区：
文献类型：
--
作者：
Kim, Jong-Seo;Dai, Ziyu;Aryal, Uma K.;Moore, Ronald J.;Camp, David G., II;Baker, Scott E.;Smith, Richard D.;Qian, Wei-Jun
已经开发了一种树脂辅助富集方法，用于特异性分离蛋白质N-末端肽，以促进蛋白水解加工（翻译后修饰的主要形式）的LC-MS/MS表征。在该方法中，蛋白质硫醇通过还原和烷基化被封闭，并且蛋白质赖氨酸残基被转化为高丝氨酸。蛋白质N-末端被选择性地转化为反应性巯基，然后含巯基的N-末端肽被具有高特异性（>97%）的巯基亲和树脂捕获。这些顺序反应的效率被证明是几乎定量的。首先将树脂辅助的N-末端肽富集方法应用于丝状真菌尼日尔曲霉的细胞裂解物。随后的C-MS/MS分析导致从690个独特蛋白质中鉴定出1672个独特蛋白质N-末端或蛋白水解切割位点。
A resin-assisted enrichment method has been developed for specific isolation of protein N-terminal peptides to facilitate LC-MS/MS characterization of proteolytic processing, a major form of posttranslational modifications. In this method, protein thiols are blocked by reduction and alkylation, and protein lysine residues are converted to homoarginines. Protein N-termini are selectively converted to reactive thiol groups, and the thiol-containing N-terminal peptides are then captured by a thiol-affinity resin with high specificity (>97%). The efficiencies of these sequential reactions were demonstrated to be nearly quantitative. The resin-assisted N-terminal peptide enrichment approach was initially applied to a cell lysate of the filamentous fungus Aspergillus niger. Subsequent C-MS/MS analyses resulted in the identification of 1672 unique protein N-termini or proteolytic cleavage sites from 690 unique proteins.
登录
查看更多内容
影响因子：
3.4
作者：
Qian, WJ;Jacobs, JM;Smith, RD
通讯作者：
Smith, RD
影响因子：
7.4
作者：
Beardsley, RL;Reilly, JP
通讯作者：
Reilly, JP
影响因子：
64.5
作者：
Mahrus S;Trinidad JC;Barkan DT;Sali A;Burlingame AL;Wells JA
通讯作者：
Wells JA
影响因子：
48
作者：
McDonald, L;Robertson, DHL;Beynon, RJ
通讯作者：
Beynon, RJ
影响因子：
7
作者：
Andersen, Mikael R.;Salazar, Margarita P.;Baker, Scott E.
通讯作者：
Baker, Scott E.