DICP OpenIR
Subject Area物理化学
Combination of online enzyme digestion with stable isotope labeling for high-throughput quantitative proteome analysis
Wang, Fangjun1; Wei, Xiaoluan1; Zhou, Hu2,3; Liu, Jing1; Figeys, Daniel2; Zou, Hanfa1; Zou HF(邹汉法)
KeywordMiscleavage Online Enzyme Digestion Quantitative Proteome Analysis Stable Isotope Labeling Technology
Source PublicationPROTEOMICS
2012-11-01
ISSN1615-9853
DOI10.1002/pmic.201200162
Volume12Issue:21Pages:3129-3137
Indexed BySCI
SubtypeArticle
Department18
Funding Project1809
Contribution Rank1, 1
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
WOS SubjectBiochemical Research Methods ; Biochemistry & Molecular Biology
WOS Research AreaBiochemistry & Molecular Biology
WOS KeywordTANDEM MASS-SPECTROMETRY ; PHASE LIQUID-CHROMATOGRAPHY ; TRYPSIN REACTOR ; COUPLED ONLINE ; CELL-CULTURE ; AMINO-ACIDS ; IDENTIFICATION ; PROTEOLYSIS ; CAPILLARY ; SILAC
AbstractVarious enzyme reactors and online enzyme digestion strategies have been developed in recent years. These reactors greatly enhanced the detection sensitivity and proteome coverage in qualitative proteomics. However, these devices have higher rates of miscleavage in protein digestion. Therefore, we investigated the effect of online enzyme digestion on the quantification accuracy of quantitative proteomics using chemical or metabolic isotope labeling approaches. The incomplete digestion would introduce some unexpected variations in comparative quantification when the samples are digested and then chemically isotope labeled in different aliquots. Even when identical protein aliquots are processed on these devices using post-digestion chemical isotope labeling and the CVs of the ratios controlled to less than 50% in replicate analyses, about 10% of the quantified proteins have a ratio greater than two-fold, whereas in theory the ratio is 1:1. Interestingly, the incomplete digestion with enzyme reactor is not a problem when metabolic isotope labeling samples were processed because the proteins are isotopically labeled in vivo prior to their simultaneous digestion within the reactor. Our results also demonstrated that both high quantification accuracy and high proteome coverage can be achieved in comparative proteome quantification using online enzyme digestion even when a limited amount of metabolic isotope labeling samples is used (1683 proteins comparatively quantified from 105 Hela cells).
Language英语
WOS IDWOS:000310564100002
Citation statistics
Cited Times:13[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://cas-ir.dicp.ac.cn/handle/321008/118443
Collection中国科学院大连化学物理研究所
Corresponding AuthorZou HF(邹汉法)
Affiliation1.Chinese Acad Sci, Dalian Inst Chem Phys, CAS Key Lab Separat Sci Analyt Chem, Natl Chromatog Res & Anal Ctr, Dalian 116023, Peoples R China
2.Univ Ottawa, Ottawa Inst Syst Biol, Ottawa, ON, Canada
3.Chinese Acad Sci, Shanghai Inst Mat Med, Shanghai 200031, Peoples R China
Recommended Citation
GB/T 7714
Wang, Fangjun,Wei, Xiaoluan,Zhou, Hu,et al. Combination of online enzyme digestion with stable isotope labeling for high-throughput quantitative proteome analysis[J]. PROTEOMICS,2012,12(21):3129-3137.
APA Wang, Fangjun.,Wei, Xiaoluan.,Zhou, Hu.,Liu, Jing.,Figeys, Daniel.,...&邹汉法.(2012).Combination of online enzyme digestion with stable isotope labeling for high-throughput quantitative proteome analysis.PROTEOMICS,12(21),3129-3137.
MLA Wang, Fangjun,et al."Combination of online enzyme digestion with stable isotope labeling for high-throughput quantitative proteome analysis".PROTEOMICS 12.21(2012):3129-3137.
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