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Fast MS/MS acquisition without dynamic exclusion enables precise and accurate quantification of proteome by MS/MS fragment intensity.

ABSTRACT: Most currently proteomic studies use data-dependent acquisition with dynamic exclusion to identify and quantify the peptides generated by the digestion of biological sample. Although dynamic exclusion permits more identifications and higher possibility to find low abundant proteins, stochastic and irreproducible precursor ion selection caused by dynamic exclusion limit the quantification capabilities, especially for MS/MS based quantification. This is because a peptide is usually triggered for fragmentation only once due to dynamic exclusion. Therefore the fragment ions used for quantification only reflect the peptide abundances at that given time point. Here, we propose a strategy of fast MS/MS acquisition without dynamic exclusion to enable precise and accurate quantification of proteome by MS/MS fragment intensity. The results showed comparable proteome identification efficiency compared to the traditional data-dependent acquisition with dynamic exclusion, better quantitative accuracy and reproducibility regardless of label-free based quantification or isobaric labeling based quantification. It provides us with new insights to fully explore the potential of modern mass spectrometers. This strategy was applied to the relative quantification of two human disease cell lines, showing great promises for quantitative proteomic applications.

SUBMITTER: Zhang S

PROVIDER: S-EPMC4873735 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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Fast MS/MS acquisition without dynamic exclusion enables precise and accurate quantification of proteome by MS/MS fragment intensity.

Zhang Shen S Wu Qi Q Shan Yichu Y Zhao Qun Q Zhao Baofeng B Weng Yejing Y Sui Zhigang Z Zhang Lihua L Zhang Yukui Y

Scientific reports 20160520

Most currently proteomic studies use data-dependent acquisition with dynamic exclusion to identify and quantify the peptides generated by the digestion of biological sample. Although dynamic exclusion permits more identifications and higher possibility to find low abundant proteins, stochastic and irreproducible precursor ion selection caused by dynamic exclusion limit the quantification capabilities, especially for MS/MS based quantification. This is because a peptide is usually triggered for f ...[more]

PMID: 27198003

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Project description:Limonoids are bioactive plant specialized metabolites found in the Meliaceae family. The basic limonoids, i.e., azadiradione, epoxyazadiradione, and gedunin have been exploited for various bioactivities and therefore are the potential drug leads for tomorrow. However, their low abundance, structural similarity, and lack of adequate mass fragmentation data have hampered their accurate identification and quantification from various sources. In the present study, basic limonoids such as azadirone, azadiradione, epoxyazadiradione, and gedunin isolated from Neem were utilized for the synthesis of their derivatives and isotopologs. A total of 30 one compounds were used in this study among which five were isolated, two were biotransformed, and 24 were synthesized. Among the synthesized compounds nine are novel compounds including six deuterated analogs/isotopologs which are (1,3-2H)-1,2-dihydro-3β-hydroxyazadiradione (9), (1,3,16-2H)-1,2-dihydro-3β-16β-dihydroxyazadiradione (10), 3β-hydroxyazadiradione (11), 3β-16β-dihydroxyazadiradione (12), (3-2H)-3β-hydroxyazadiradione (13), (3,16-2H)-3β-16β-dihydroxyazadiradione (14), (1,3,7-2H)-1,2-dihydro-3β-hydroxy-7-deacetylazadiradione (15), 1,2,20,21,22,23-hexahydroazadiradione (17), and (1,3-2H)-1,2-dihydro-3β-hydroxygedunin (29). These limonoids along with their semisynthesized derivatives were subjected to ultra high performance liquid chromatography mass spectrometry (UHPLC-MS/MS) and the fragmentation pathway was established based on structure-fragment relationships (SFRs), utilizing high resolution MS/MS data. We have developed a most reliable and easily reproducible protocol describing in depth analysis of SFRs based on the structural modifications and synthesis of isotopologs. Also, the MS/MS fragment library of these basic limonoids generated in this study acts as a fingerprint for accurate identification and quantification of limonoids by MS/MS analysis in various plant tissue extracts, phytopharmaceutical formulations and biological samples.

Dataset Information

Fast MS/MS acquisition without dynamic exclusion enables precise and accurate quantification of proteome by MS/MS fragment intensity.

Publications

Fast MS/MS acquisition without dynamic exclusion enables precise and accurate quantification of proteome by MS/MS fragment intensity.

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