镁离子对碱性磷酸酶折叠过程的影响

首都医科大学学报 ›› 2003, Vol. 24 ›› Issue (3): 221-224.

镁离子对碱性磷酸酶折叠过程的影响

宋晓红¹, 闫淑莲¹, 田晓娟¹, 张英侠¹, 周海梦²

1. 首都医科大学基础医学院化学教研室;2. 清华大学生物科学与技术系

收稿日期:2003-03-12 修回日期:1900-01-01 出版日期:2003-07-15 发布日期:2003-07-15

Effects of Magnesium Ions on Refolding of Calf Intestinal Alkaline Phosphatase

Song Xiaohong¹, Yan Shulian¹, Tian Xiaojuan¹, Zhang Yingxia¹, Zhou Haimeng²

1. Department of Chemistry, School of Basic Sciences, Capital University of Medical Sciences;2. Department of Biological Sciences & Technology,Tsinghua University

Received:2003-03-12 Revised:1900-01-01 Online:2003-07-15 Published:2003-07-15

摘要/Abstract

摘要： 对小牛肠碱性磷酸酶再折叠的机制及其影响因素进行了探讨。在25℃室温下,以3mol/L盐酸胍对小牛肠碱性磷酸酶(CIP)进行了2h的去折叠实验,然后再以Tris-HCl缓冲液对变性体系进行20倍稀释,观察该酶活力恢复的动力学过程并与镁离子参与下的稀释复性过程进行比较。并就不同实验条件下含镁稀释或不含镁稀释,获得的最终酶活力、酶活力恢复水平、活力恢复速率常数及数据的相对标准偏差等指标进行了探讨。认为镁是CIP位点的特异性“效应分子”,它与酶中有关基团的配位,稳定了酶活性部位的构象,使得自发折叠过程能够向类天然态的方向进行。

关键词: 碱性磷酸酶, 再折叠, 镁

Abstract: Recently the refolding studies have been concentrated on alkaline phosphatase from E. Coli. The present experiments studied the refolding of mammalian enzyme. Calf intestinal alkaline phosphatase (CIP) was denatured in 3 mol/Lguanidine hydrochloride solutions for 2 h at 25 ℃. Under suitable renaturation conditions, the kinetic courses of enzymatic activity recovery were investigated in the absence and presence of magnesium ions at 2 mmol/Lconcentration, and compared to each other. The two remarkably different courses characterized by the final values of activity recovery, recovery levels, kinetic constants of activity recovery and relative standard deviation of the data indicated that magnesium ions behaved like "site-specific" effectors which were bound to the concerning residues of enzyme protein and stabilized the conformation of active site domain, thus guided the refolding process in a productive pathway.

Key words: alkaline phosphatase, refolding, magnesium

中图分类号:

Q556

宋晓红;闫淑莲;田晓娟;张英侠;周海梦. 镁离子对碱性磷酸酶折叠过程的影响[J]. 首都医科大学学报, 2003, 24(3): 221-224.

Song Xiaohong;Yan Shulian;Tian Xiaojuan;Zhang Yingxia;Zhou Haimeng. Effects of Magnesium Ions on Refolding of Calf Intestinal Alkaline Phosphatase[J]. Journal of Capital Medical University, 2003, 24(3): 221-224.

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