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学科主题: 物理化学
题名: Biphasic Microreactor for Efficient Membrane Protein Pretreatment with a Combination of Formic Acid Assisted Solubilization, On-Column pH Adjustment, Reduction, Alkylation, and Tryptic Digestion
作者: Zhao, Qun1, 2;  Liang, Yu1;  Yuan, Huiming1;  Sui, Zhigang1;  Wu, Qi1, 2;  Liang, Zhen1;  Zhang, Lihua1;  Zhang, Yukui1
通讯作者: 张丽华
刊名: ANALYTICAL CHEMISTRY
发表日期: 2013-09-17
DOI: 10.1021/ac402076u
卷: 85, 期:18, 页:8507-8512
收录类别: SCI
合作性质: 
文章类型: Article
部门归属: 18
项目归属: 1810
产权排名: 待补充
WOS标题词: Science & Technology ;  Physical Sciences
类目[WOS]: Chemistry, Analytical
资助者: 1,1
研究领域[WOS]: Chemistry
英文摘要: Combining good dissolving ability of formic acid (FA) for membrane proteins and excellent complementary retention behavior of proteins on strong cation exchange (SCX) and strong anion exchange (SAX) materials, a biphasic microreactor was established to pretreat membrane proteins at microgram and even nanogram levels. With membrane proteins solubilized by FA, all of the proteomics sample processing procedures, including protein preconcentration, pH adjustment, reduction, and alkylation, as well as tryptic digestion, were integrated into an "SCX-SAX" biphasic capillary column. To evaluate the performance of the developed microreactor, a mixture of bovine serum albumin, myoglobin, and cytochrome c was pretreated. Compared with the results obtained by the traditional in-solution process, the peptide recovery (93% vs 83%) and analysis throughput (3.5 vs 14 h) were obviously improved. The microreactor was further applied for the pretreatment of 14 mu g of membrane proteins extracted from rat cerebellums, and 416 integral membrane proteins (IMPs) (43% of total protein groups) and 103 transmembrane peptides were identified by two-dimensional nanoliquid chromatography-electrospray ionization tandem mass spectrometry (2D nano-LC-ESI-MS/MS) in triplicate analysis. With the starting sample preparation amount decreased to as low as 50 ng, 64 IMPs and 17 transmembrane peptides were identified confidently, while those obtained by the traditional in-solution method were 10 and 1, respectively. All these results demonstrated that such an "SCX-SAX" based biphasic microreactor could offer a promising tool for the pretreatment of trace membrane proteins with high efficiency and throughput.
关键词[WOS]: TANDEM MASS-SPECTROMETRY ;  STRONG-CATION-EXCHANGE ;  PHASE LIQUID-CHROMATOGRAPHY ;  INTEGRAL MEMBRANE ;  PROTEOMIC ANALYSIS ;  IDENTIFICATION TECHNOLOGY ;  SHOTGUN PROTEOMICS ;  TRYPSIN DIGESTION ;  BRAIN PROTEOME ;  METHANOL
语种: 英语
原文出处: 查看原文
WOS记录号: WOS:000330017400007
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/119507
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, Key Lab Separat Sci Analyt Chem, Natl Chromatog R&A Ctr, Dalian 116023, Peoples R China
2.Univ Chinese Acad Sci, Beijing 100039, Peoples R China

Recommended Citation:
Zhao, Qun,Liang, Yu,Yuan, Huiming,et al. Biphasic Microreactor for Efficient Membrane Protein Pretreatment with a Combination of Formic Acid Assisted Solubilization, On-Column pH Adjustment, Reduction, Alkylation, and Tryptic Digestion[J]. ANALYTICAL CHEMISTRY,2013,85(18):8507-8512.
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