多层开管毛细管酶反应器的制备及在蛋白质酶切中的应用

doi:10.3969/j.issn.0251-0790.2012.02.010

高等学校化学学报 ›› 2012, Vol. 33 ›› Issue (02): 268.doi: 10.3969/j.issn.0251-0790.2012.02.010

多层开管毛细管酶反应器的制备及在蛋白质酶切中的应用

胡佳薇¹, 申烨华¹, 秦伟捷², 张养军², 钱小红²

1. 西北大学合成与天然功能分子化学教育部重点实验室, 新药研究所, 化学与材料科学学院, 西安 710069;
2. 蛋白质组学国家重点实验室, 北京蛋白质组研究中心, 北京放射与辐射医学研究所, 北京 102206

收稿日期:2011-02-07 出版日期:2012-02-10 发布日期:2012-01-13
通讯作者: 钱小红, 女, 博士, 研究员, 博士生导师, 主要从事蛋白质组学研究. E-mail: qianxh1@yahoo.com.cn 张养军, 男, 博士, 副研究员, 主要从事蛋白质组学新方法和新技术研究. E-mail: zhangyangjun6314@yahoo.com.cn E-mail:qianxh1@yahoo.com.cn;zhangyangjun6314@yahoo.com.cn
基金资助:
国家"九七三"计划项目(批准号: 2010CB912701, 2012CB910603)和国家自然科学基金(批准号: 30621063, 20735005, 20875101, 20905077, 20975059)资助.

Preparation of Multi-layer Open Capillary Immobilized Enzyme Reactor and Its Application in Digestion of Proteins

HU Jia-Wei¹, SHEN Ye-Hua¹, QIN Wei-Jie², ZHANG Yang-Jun², Qian Xiao-Hong²

1. Key Laboratory of Synthetic and Natural Functional Molecule Chemistry of Ministry of Education, Drug Research Institute, College of Chemistry and Materials Science, Northwest University, Xi'an 710069, China;
2. State Key Laboratory of Proteomics, Beijing Proteome Research Center, Beijing Institute of Radiation Medicine, Beijing 102206, China

Received:2011-02-07 Online:2012-02-10 Published:2012-01-13

摘要/Abstract

摘要： 以石英毛细管作为酶固定化的载体, 在毛细管内壁上逐步合成树枝形大分子聚酰胺-胺(PAMAM), 再通过交联剂戊二醛将胰蛋白酶直接键合到该大分子的末端氨基上, 并对酶固定化条件进行了优化, 制备了多层酶反应器. 利用该酶反应器对马心细胞色素C等蛋白质进行了酶切, 并对酶切的条件进行了优化. 实验结果表明, 该固定化酶反应器具有较高的酶切效率、良好的重现性和稳定性, 可用于蛋白质组学的研究.

关键词: 开管柱, 固定酶反应器, 蛋白质酶切, 树枝形大分子

Abstract: With the capillary as the carrier of enzyme immobilization, poly(amidoamine) was gradually bon-ded to the inner wall of the capillary, then the trypsin was attached at the amino group of poly(amidoamine) with glutaraldehyde, and finally the multi-layer open capillary immobilized enzyme reactor was prepared after enzyme immobilization conditions had been optimized. Cyt-c was chosen as a model protein for evaluation of the performance of the multi-layer open capillary immobilized enzyme reactor, and its digestion conditions were optimized. The results demonstrate that the multi-layer immobilized enzyme reactor has high efficiency, good reproducibility and stability, and can be further applied in future proteomics study.

Key words: Open-tubular capillary, Immobilized enzyme reactor, Protein digestion, Dendritical macromolecule

中图分类号:

O657

TrendMD:

胡佳薇, 申烨华, 秦伟捷, 张养军, 钱小红. 多层开管毛细管酶反应器的制备及在蛋白质酶切中的应用. 高等学校化学学报, 2012, 33(02): 268.

HU Jia-Wei, SHEN Ye-Hua, QIN Wei-Jie, ZHANG Yang-Jun, Qian Xiao-Hong. Preparation of Multi-layer Open Capillary Immobilized Enzyme Reactor and Its Application in Digestion of Proteins. Chem. J. Chinese Universities, 2012, 33(02): 268.

参考文献

[1] Mann M., Hojrup P., Roepstorff P.. Biol. Mass. Spectrom.[J], 1993, 22(6): 338-345 [2] Yates J. R. 3rd., Speicher S., Griffin P. R., Hunkapiller T.. Anal. Biochem.[J], 1993, 214(2): 397-408 [3] Mann M., Wilm M.. Anal. Chem.[J], 1994, 66(24): 4390-4399 [4] Ma J. F., Liang Z., Qiao X. Q., Deng Q. L., Tao D. Y., Zhang L. H., Zhang Y. K.. Anal. Chem.[J], 2008, 80: 2949-2956 [5] Freije J. R., Mulder P. P., Werkman W., Rieux L., Niederlander H. A. G., Verpoorte E., Bischoff R.. J. Proteome Res.[J], 2005, 4: 1805-1813 [6] Duan J. C., Liang Z., Yang C., Zhang L., Zhang W., Zhang Y.. Proteomics[J], 2006, 6: 412-419 [7] Shui W., Fan J., Yang P., Zhai J., Lei J., Yan Y., Zhao D., Chen X.. Anal. Chem.[J], 2006, 78(14): 4811-4819 [8] Li Y., Yan B., Deng C., Tang J., Liu J. Y., Zhang X. M.. Proteomics[J], 2007, 7: 3661-3671 [9] Liu Y., Xue Y., Ji J., Chen X., Kong J. L., Yang P. Y., Girault H. H., Liu B. H.. Molecular & Cellular Proteomics[J], 2007, 6: 1428-1436 [10] Gao J., Xu J. D., Locascio L. E., Lee C. S.. Anal. Chem.[J], 2001, 73: 2648-2655 [11] Cooper J. W., Chen J., Li Y., Lee C. S.. Anal. Chem.[J], 2003, 75: 1067-1074 [12] Liu Y., Lu H. J., Zhong W., Kong J. L., Yang P. Y., Girault H. H., Liu B. H.. Anal.Chem. [J], 2006, 78: 801-808 [13] Calleri E., Temporini C., Perani E., De P. A., Lubda D., Mellerio G., Sala A., Galliano M., Caccialanza G., Massolini G.. J. Proteome Res.[J], 2005, 4(2): 481-490 [14] Calleri E., Temporini C., Perani E., Stella C., Rudaz S., Lubda D., Mellerio G., Veuthey J. L., Caccialanza G., Massolini G.. J. Chromatogr. A[J], 2004, 1045: 99-109 [15] Ye M. L., Hu S., Schoenherr R. M., Dovichi N. J.. Electrophoresis[J], 2004, 25(9): 1319-1326 [16] Amankwa L. N., Kuhr W. G.. Anal. Chem.[J], 1992, 64: 1610-1613 [17] Bossi A., Guizzardi L., D'Acunto M. R., Righetti P. G.. Anal. Bioanal. Chem.[J], 2004, 378: 1722-1728 [18] Hsieh Y. L. F., Wang H. Q., Elicone C., Mark J., Martin S. A., Regnier F.. Anal. Chem.[J], 1996, 68: 455-462 [19] Slysz G. W., Schriemer D. C.. Anal. Chem.[J], 2005, 77: 1572-1579 [20] Slysz G. W., Lewis D. F., Schriemer D. C.. J. Proteome Res.[J], 2006, 5: 1959-1966 [21] GUO Zhong(郭忠), ZHANG Qing-Chun(张清春), LEI Zheng-Deng(雷政登), KONG Liang(孔亮), MAO Xi-Qin(毛希琴), ZOU Han-Fa(邹汉法). Chem. J. Chinese Universities(高等学校化学学报)[J], 2002, 23(7): 1277-1280 [22] ZHANG Zhi-Sheng(张志胜), YANG Yi(杨屹). Chem. J. Chinese Universities(高等学校化学学报)[J], 2006, 27(1): 47-51 [23] Mckeehan W. L.. Cell Biology International Reports[J], 1977, 1(4): 335-343 [24] CHEN Min-Heng(陈敏恒), CONG De-Zi(丛德滋), FANG Tu-Nan(方图南), QI Ming-Zhai(齐鸣斋). Principle of Chemical Engineering(化工原理)[M], Beijing: Chemical Industry Press, 1999: 38-47

多层开管毛细管酶反应器的制备及在蛋白质酶切中的应用

Preparation of Multi-layer Open Capillary Immobilized Enzyme Reactor and Its Application in Digestion of Proteins

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