mTRAQ-based quantitative analysis combined with peptide fractionation based on cysteinyl peptide enrichment
- Authors
- Yeom, Jeonghun; Kang, Min Jung; Shin, Dongyun; Song, Hyun Kyu; Lee, Cheolju; Lee, Ji Eun
- Issue Date
- 15-May-2015
- Publisher
- ACADEMIC PRESS INC ELSEVIER SCIENCE
- Keywords
- Quantification; Peptide fractionation; Cysteinyl peptide enrichment; Mass spectrometry
- Citation
- ANALYTICAL BIOCHEMISTRY, v.477, pp.41 - 49
- Journal Title
- ANALYTICAL BIOCHEMISTRY
- Volume
- 477
- Start Page
- 41
- End Page
- 49
- URI
- https://scholarworks.bwise.kr/gachon/handle/2020.sw.gachon/10527
- DOI
- 10.1016/j.ab.2015.03.005
- ISSN
- 0003-2697
- Abstract
- In the present study, the fractionation scheme for cysteinyl peptide enrichment (CPE) was combined with the mass differential tags for relative and absolute quantification (mTRAQ) method to reduce sample complexity and increase proteome coverage. Cysteine residues of the proteins were first alkylated using iodoacetyl PEG(2)-biotin instead of other conventional alkylating agents such as iodoacetamide. After trypsin digestion, amine groups were labeled with mTRAQ and these labeled peptides were fractionated according to the presence or absence of cysteine residues using avidin-biotin affinity chromatography. With these approaches, we were able to divide the peptides into the two fractions with more than 90% fractionation efficiency for standard protein and MCF7 cell lysate. When the fractionation strategy was applied to colorectal cancer tissue samples, we were able to obtain quantitative information that was consistent with the previous study based on mTRAQ quantification, implying that the cysteine-based fractionation method does not affect mTRAQ quantification. We expect that the mTRAQ-based quantitative analysis combined with peptide fractionation through the CPE strategy would allow for deep-down analysis of proteome samples and ultimately for increasing proteome coverage with simultaneous quantification for biomarker discovery. (C) 2015 Elsevier Inc. All rights reserved.
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