期刊
RAPID COMMUNICATIONS IN MASS SPECTROMETRY
卷 31, 期 2, 页码 171-178出版社
WILEY-BLACKWELL
DOI: 10.1002/rcm.7779
关键词
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资金
- Natural Science Foundation of Zhejiang Province [LY15H160004]
- third level in Zhejiang Province 151 talents project
- Public Projects of Zhejiang Province [2015C37064]
- Zhejiang Provincial Natural Science Foundation of China [LY16H160035]
- Zhejiang Province TCM Science and Technology plan [2016ZB002]
RationaleMass spectrometry (MS)-based protein identification depends mainly on protein extraction and digestion. Although sodium dodecyl sulfate (SDS) can preclude enzymatic digestion and interfere with MS analysis, it is still the most widely used surfactant in these steps. To overcome these disadvantages, a SDS-compatible proteomic technique for SDS removal prior to MS-based analyses was developed, namely filter-aided sample preparation (FASP). MethodsHerein, based on the effectiveness of sodium deoxycholate and a detergent removal spin column, we developed a modified FASP (mFASP) method and compared its overall performance, total number of peptides and proteins identified for shotgun proteomic experiments with that of the FASP method. ResultsIdentification of 4570392 and 9139 +/- 317 peptides and description of 862 +/- 46 and 1377 +/- 33 protein groups with two or more peptides from the ovarian cancer cell line A2780 was accomplished by FASP and mFASP methods, respectively. The mFASP method (21.2 +/- 0.2%) had higher average peptide to protein coverage than FASP method (13.2 +/- 0.5%). More hydrophobic peptides were identified by mFASP than by FASP, as indicated by the GRAVY score distribution. ConclusionsThe reported method enables reliable and efficient identification of proteins and peptides in whole-cell extracts containing SDS. The new approach allows for higher throughput (the simultaneous identification of more proteins), a more comprehensive investigation of proteins, and potentially the discovery of new biomarkers. Copyright (c) 2016 John Wiley & Sons, Ltd.
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