Method of Detecting Phospholipase D Activity and Analyses of Kinetic Parameters in Chorispora bungean

doi:10.11733/j.issn.1007-0435.2014.02.025

›› 2014, Vol. 22 ›› Issue (2): 375-379.DOI: 10.11733/j.issn.1007-0435.2014.02.025

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Method of Detecting Phospholipase D Activity and Analyses of Kinetic Parameters in Chorispora bungean

YANG Ning¹, WANG Cheng-liang¹, CHEN Xia¹, DING Fang-xia¹, LI Yi-kun¹, ZHAO Cong-cong¹, An Li-zhe²

1. College of Life Science, Northwest Normal University, Lanzhou, Gansu Province 730070, China;
2. College of Life Science, Lanzhou University, Lanzhou, Gansu Province 730070, China

Received:2013-06-11 Revised:2013-09-17 Online:2014-04-15 Published:2014-04-21

高山离子芥磷脂酶D活性测定及酶动力学分析

杨宁¹, 王程亮¹, 陈霞¹, 丁芳霞¹, 李宜珅¹, 赵聪聪¹, 安黎哲²

1. 西北师范大学生命科学学院, 甘肃兰州 730070;
2. 兰州大学生命科学学院, 甘肃兰州 730000

通讯作者: 安黎哲
作者简介:杨宁（1973-），女，山东青州人，博士，副教授，主要从事植物分子细胞生物学研究，E-mail：xbsd-yn@163.com
基金资助:
国家自然科学基金（31160087）（31360061）；甘肃省财政厅高校基本科研业务费项目；甘肃省教育厅基金（1101-06）；西北师范大学学生学术科研资助项目资助

Abstract

Abstract: Chorispora bungeana is a representative alpine-subnival herbaceous plant with special molecular and physiological mechanism response to abiotic stresses. Phospholipase D (PLD) is not only a major kind of phospholipid hydrolases in plant, but also an important signal transduction enzyme. Using enzyme couple colorimetric method, the better reaction system is studied. The proper content of membrane protein is 35 μg in 200 μL reaction system, the reaction time is 30 min at 25℃ and the suitable pH is 7.0. Further analyses of Chorispora bungean callus from different fractions show that the different binding states of PLD have different kinetic parameters. These results establish the research foundation of PLD responding to environmental stress.

Key words: Chorispora bungean, Phospholipase D, Activity, Detection method

摘要： 高山离子芥（Chorispora bungean）是一种高抗逆的多年生高山草本植物，具有特殊的分子及生理抗逆响应机制。磷脂水解酶磷脂酶D （PLD）是重要的跨膜信号转导酶。以高山离子芥愈伤组织为试材，利用酶联免疫分光光度计法，优化磷脂酶D活性测定体系，建立了在25℃反应温度条件下，200 μL反应体系中，膜蛋白的含量35 μg、反应时间30 min、pH为7.0的缓冲液系统最适合高山离子芥愈伤组织磷脂酶D活性测定体系，分析表明不同膜结合态磷脂酶D的酶动力学特征存在差异，为进一步探讨磷脂酶D在高山离子芥响应逆境胁迫中的作用奠定基础。

关键词: 高山离子芥, 磷脂酶D, 活性, 测定方法

CLC Number:

YANG Ning, WANG Cheng-liang, CHEN Xia, DING Fang-xia, LI Yi-kun, ZHAO Cong-cong, An Li-zhe. Method of Detecting Phospholipase D Activity and Analyses of Kinetic Parameters in Chorispora bungean[J]. , 2014, 22(2): 375-379.

杨宁, 王程亮, 陈霞, 丁芳霞, 李宜珅, 赵聪聪, 安黎哲. 高山离子芥磷脂酶D活性测定及酶动力学分析[J]. , 2014, 22(2): 375-379.

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Method of Detecting Phospholipase D Activity and Analyses of Kinetic Parameters in Chorispora bungean

高山离子芥磷脂酶D活性测定及酶动力学分析

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