Top-down analysis of basic proteins by microchip capillary electrophoresis mass spectrometry
Top-down analysis of basic proteins by microchip capillary electrophoresis mass spectrometry
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DOI：
10.1002/rcm.2541
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发表时间：
2006-01-01
影响因子：
2
通讯作者：
Nishimura, Yoshifumi
中科院分区：
文献类型：
--
作者：
Akashi, Satoko;Suzuki, Koichi;Nishimura, Yoshifumi
建立了一种用于蛋白质快速鉴定的微芯片毛细管电泳/电喷雾电离质谱（mu chip-CE/ESI-MS）系统。毛细管电泳（CE）能够以高分辨率快速分析极少量的样品，例如在飞摩尔水平。通过将普通毛细管缩小到微芯片毛细管，期望更快的CE/MS分析。虽然CE分离的优点是快速和高分辨率，但由于蛋白质与微通道表面相互作用引起的电渗透流（EOF）不稳定性导致中性pH条件下碱性蛋白质分析的重现性较低。采用四极杆(Q)和飞行时间（TOF）混合仪器，在微通道表面包被碱性聚合物聚-323，可以分离和检测pI值大于7.5的碱性蛋白。通过增加芯片- ce /ESI-MS分析小蛋白时的锥电压和碰撞电压，还可以获得一些含有序列信息的产物离子，即通过芯片- ce /MS系统可以实现对蛋白质的“自上而下”分析。据我们所知，这是第一个用mu chip-CE/MS对蛋白质进行“自上而下”分析的报告。与传统的液相色谱(LC)/ESI-MS相比，mu chip-CE/MS所需的分析时间更短，样本量更少，因此mu chip-CE/MS有望用于蛋白质的高通量表征。版权所有(c) 2006约翰威利父子有限公司
A system of microchip capillary electrophoresis/electrospray ionization mass spectrometry (mu chip-CE/ESI-MS) for rapid characterization of proteins has been developed. Capillary electrophoresis (CE) enables rapid analysis of a sample present in very small quantity, such as at femtomole levels, at high resolution. Faster CE/MS analysis is expected by downsizing the normal capillary to the microchip (mu chip) capillary. Although rapidity and high resolution are advantages of CE separation, electro-osmotic flow (EOF) instability caused by the interaction between proteins and the microchannel surface results in low reproducibility in the analysis of basic proteins under neutral pH conditions. By coating the microchannel surface with a basic polymer, polyE-323, basic proteins, which have pI values of over 7.5, could be separated and detected by mu chip-CE/MS on quadrupole (Q) and time-of-flight (TOF) hybrid instruments. By increasing the cone and collision voltages during the analysis by mu chip-CE/ESI-MS of a small protein, some product ions, which contain the sequence information, could also be obtained, i.e., 'top-down' analysis of the protein could be accomplished with this mu chip-CE/MS system. To our knowledge, this is the first report of 'top-down' analysis of a protein by mu chip-CE/MS. Since it requires a much shorter time and a smaller sample amount for analysis than the conventional liquid chromatography (LC)/ESI-MS method, mu chip-CE/MS promises to be suitable for the high-throughput characterization of proteins. Copyright (c) 2006 John Wiley & Sons, Ltd.