Rigorous Determination of the Stoichiometry of Protein Phosphorylation Using Mass Spectrometry

被引：35

作者：

Johnson, Hannah ^{[1
]}

Eyers, Claire E. ^{[1
]}

Eyers, Patrick A. ^{[2
]}

Beynon, Robert J. ^{[3
]}

Gaskell, Simon J. ^{[1
]}

机构：

[1] Univ Manchester, Manchester Interdisciplinary Bioctr, Sch Chem, Michael Barber Ctr Mass Spectrometry, Manchester M1 7DN, Lancs, England

[2] Univ Sheffield, Yorkshire Canc Res Inst Canc Studies, Sheffield, S Yorkshire, England

[3] Univ Liverpool, Fac Vet Sci, Prote & Funct Genom Grp, Liverpool L69 3BX, Merseyside, England

来源：

JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY | 2009年 / 20卷 / 12期

基金：

英国工程与自然科学研究理事会; 英国生物技术与生命科学研究理事会; 英国医学研究理事会;

关键词：

MULTIPLEXED ABSOLUTE QUANTIFICATION; CONCATENATED SIGNATURE PEPTIDES; LABELING STRATEGY; ACTIVATION LOOP; KINASE; SITE; IDENTIFICATION; CHROMATOGRAPHY; MPS1; DYNAMICS;

D O I：

10.1016/j.jasms.2009.08.009

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Quantification of the stoichiometry of phosphorylation is usually achieved using a mixture of phosphatase treatment and differential isotopic labeling. Here, we introduce a new approach to the concomitant determination of absolute protein concentration and the stoichiometry of phosphorylation at predefined sites. The method exploits QconCAT to quantify levels of phosphorylated and nonphosphorylated peptide sequences in a phosphoprotein. The nonphosphorylated sequence is used to determine the absolute protein quantity and serves as a reference to calculate the extent of phosphorylation at the second peptide. Thus, the stoichiometry of phosphorylation and the absolute protein concentration can be determined accurately in a single experiment. (J Am Soc Mass Spectrom 2009, 20, 2211-2220) (C) 2009 American Society for Mass Spectrometry

引用

页码：2211 / 2220

页数：10

共 23 条

[1] Multiplexed absolute quantification in proteomics using artificial QCAT proteins of concatenated signature peptides [J].