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题名: Determination of CK2 Specificity and Substrates by Proteome-Derived Peptide Libraries
作者: Wang, Chunli1, 2;  Ye, Mingliang1;  Bian, Yangyang1, 2;  Liu, Fangjie1, 2;  Cheng, Kai1, 2;  Dong, Mingming1, 2;  Dong, Jing1;  Zou, Hanfa1
关键词: peptide library ;  casein kinase 2 ;  specificity ;  substrates ;  quantitative phosphoproteomics
刊名: JOURNAL OF PROTEOME RESEARCH
发表日期: 2013-08-01
DOI: 10.1021/pr4002965
卷: 12, 期:8, 页:3813-3821
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology ;  Life Sciences & Biomedicine
类目[WOS]: Biochemical Research Methods
研究领域[WOS]: Biochemistry & Molecular Biology
英文摘要: Understanding the specificity of kinases enables prediction of their substrates and uncovering kinase functions in signaling pathways. Traditionally synthesized peptide libraries are used to determine the kinase specificity. In this study, a proteomics-based method was developed to determine the specificity of kinase by taking the advantages of proteome-derived peptide libraries and quantitative proteomics. Proteome-derived peptide libraries were constructed by digesting proteins in total cell lysate followed with dephosphorylation of the resulting peptides. After incubating the peptide libraries with/without CK2 for in vitro kinase assay, stable isotopic labeling based quantitative phosphoproteomics was applied to distinguish the in vitro phosphosites generated by CK2. By using the above approach, 404 CK2 in vitro phosphosites were identified by ID LC-MS/MS. Those sites allowed the statistic determination of the CK2 specificity. In addition to the easy construction of the proteome-derived peptide library, another significant advantage of this method over the method with synthesized peptide libraries is that the identified phosphosites could be directly mapped to proteins for the screening of putative kinase substrates. It was found that the confidence for substrate identification could be significantly improved by comparing the in vitro CK2 sites with the in vivo sites identified by phosphoproteomics analysis of the same cell lines. By applying this integrated strategy, 138 phosphosites from 105 putative CK2 substrates of high confidence were determined.
关键词[WOS]: ION AFFINITY-CHROMATOGRAPHY ;  KINASE CK2 ;  PHOSPHOPROTEOME ANALYSIS ;  MASS-SPECTROMETRY ;  PHOSPHORYLATION ;  ENRICHMENT ;  LEUKEMIA ;  WEBLOGO ;  TARGET
语种: 英语
WOS记录号: WOS:000322852800025
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/137868
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

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作者单位: 1.Chinese Acad Sci, Dalian Inst Chem Phys, Key Lab Separat Sci Analyt Chem, Natl Chromatog R&A Ctr, Dalian 116023, Peoples R China
2.Chinese Acad Sci, Grad Sch, Beijing 100049, Peoples R China

Recommended Citation:
Wang, Chunli,Ye, Mingliang,Bian, Yangyang,et al. Determination of CK2 Specificity and Substrates by Proteome-Derived Peptide Libraries[J]. JOURNAL OF PROTEOME RESEARCH,2013,12(8):3813-3821.
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