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学科主题物理化学
A rapid protein sample preparation method based on organic-aqueous microwave irradiation technique
Wu, Shuaibin1,2; Zhang, Hongfei2; Yang, Kaiguang1; Ma, Junfeng3; Liang, Zhen1; Zhang, Lihua1; Zhang, Yukui1; Wu SB(吴帅宾)
关键词Maldi-tof Ms Microwave Organic-aqueous Solvent Protein Identification Digestion
刊名SCIENCE CHINA-CHEMISTRY
2015-03-01
DOI10.1007/s11426-014-5163-2
58期:3页:526-531
收录类别SCI
文章类型Article
WOS标题词Science & Technology ; Physical Sciences
类目[WOS]Chemistry, Multidisciplinary
研究领域[WOS]Chemistry
关键词[WOS]PHASE LIQUID-CHROMATOGRAPHY ; TANDEM MASS-SPECTROMETRY ; MALDI-TOF-MS ; SOLVENT SYSTEMS ; EFFICIENT PROTEOLYSIS ; ENZYMATIC DIGESTION ; SILICA MICROSPHERES ; FORMIC-ACID ; IDENTIFICATION ; PROTEOMICS
英文摘要Fast and efficient sample preparation methods are a prerequisite for protein identification in bottom-up proteomics. Here, an innovative microwave irradiation sample preparation method was developed based on an optimized organic-aqueous solvent system for protein identification. Specifically, protein solutions containing high-concentration acetonitrile were subjected to 5 min microwave irradiation. After cooling down, trypsin was added and the digestion was performed with 30 s microwave irradiation, and the resulting peptides were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A shortened processing time of only 5.5 min is needed with this method (more than 12 h is necessary in the traditional overnight protein sample preparation). Moreover, due to the absence of urea and other chaotropic reagents, the digests can be readily identified by MALDI-TOF MS. When an assessment of this method was performed by digesting a model protein BSA, 69% +/- 3% sequence coverage corresponding to 47 +/- 3 peptides was obtained, which shows better protein identification than that from the standard overnight protein sample preparation method (51% +/- 2% sequence coverage and 23 +/- 1 peptides). Another model protein a-casein was used for the analysis of protein phosphorylation with the newly developed method that yielded 4 phosphopeptides with 8 phosphorylation sites, whereas 3 phosphopeptides with 2 phosphorylation sites were obtained from the traditional overnight approach. Moreover, the organic-aqueous microwave irradiation method provides effective digestion for proteins down to fmol.
语种英语
WOS记录号WOS:000350336500025
引用统计
文献类型期刊论文
条目标识符http://cas-ir.dicp.ac.cn/handle/321008/143707
专题中国科学院大连化学物理研究所
通讯作者Wu SB(吴帅宾)
作者单位1.Chinese Acad Sci, Dalian Inst Chem Phys, Natl Chromatog R&A Ctr, Key Lab Separat Sci Analyt Chem, Dalian 116023, Peoples R China
2.China Natl Tobacco Qual Supervis & Test Ctr, Zhengzhou 450001, Peoples R China
3.Johns Hopkins Univ, Sch Med, Dept Biol Chem, Baltimore, MD 21205 USA
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GB/T 7714
Wu, Shuaibin,Zhang, Hongfei,Yang, Kaiguang,et al. A rapid protein sample preparation method based on organic-aqueous microwave irradiation technique[J]. SCIENCE CHINA-CHEMISTRY,2015,58(3):526-531.
APA Wu, Shuaibin.,Zhang, Hongfei.,Yang, Kaiguang.,Ma, Junfeng.,Liang, Zhen.,...&吴帅宾.(2015).A rapid protein sample preparation method based on organic-aqueous microwave irradiation technique.SCIENCE CHINA-CHEMISTRY,58(3),526-531.
MLA Wu, Shuaibin,et al."A rapid protein sample preparation method based on organic-aqueous microwave irradiation technique".SCIENCE CHINA-CHEMISTRY 58.3(2015):526-531.
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