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学科主题: 物理化学
题名: A rapid protein sample preparation method based on organic-aqueous microwave irradiation technique
作者: Wu, Shuaibin1, 2;  Zhang, Hongfei2;  Yang, Kaiguang1;  Ma, Junfeng3;  Liang, Zhen1;  Zhang, Lihua1;  Zhang, Yukui1
通讯作者: 吴帅宾
关键词: MALDI-TOF MS ;  microwave ;  organic-aqueous solvent ;  protein identification ;  digestion
刊名: SCIENCE CHINA-CHEMISTRY
发表日期: 2015-03-01
DOI: 10.1007/s11426-014-5163-2
卷: 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
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.
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
语种: 英语
WOS记录号: WOS:000350336500025
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/143707
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

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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

Recommended Citation:
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.
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