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玉米花粉32 kD蛋白的纯化及性质

王建华, 吴英杰, 吴显荣

北京农业大学生物学院.北京 100094；中科院生物物理研究所生物大分子国家重点实验室.北京 100101

摘要

利用丙酮粉、硫酸铵分段盐析及离子交换层析技术,从玉米花粉细胞质中分离纯化了一种32 kD的可溶性蛋白,其GTPase活性大于ATPase。SDS-PAGE表明达银染电泳纯。等电聚焦电泳测得等电点为5.25。免疫印迹鉴定表明该蛋白与抗牛脑动蛋白或动力蛋白的抗体无免疫交叉反应。最大紫外吸收波长为282nm,CD谱分析说明具有球蛋白特征。

关键词： 玉米花粉; 低分子量ATPase; 提纯; 性质

Purification of 32 kD protein from maize pollen and study on some of its properties

Wang Jianhua, Wu Yingjie, Wu Xianrong

College of Biological Sciences, Beijing Agricultural University. Beijing 100094； China

Abstract

A novel 32 kD protein was purified from the cytoplasmic extracts of maize pollen by the preparation of acetone powder, ammonium sulfate fractionation, followed by DEAE-Sephadex A50 and FPLC ion-exchange chromatography. The final product of this procedure showed a single band (silver staining) on the SDS-PAGE. The purification fold was 84.5. The molecular weight of its subunit was about 32 kD as determined by SDS-PAGE and the isoelectric point was pH 5.25 by IEF-PAGE. Western-blot analysis showed that the 32 kD protein did not have specific immuno-reactions with anti-kinesin and anti-dynamin polyclonal antibodies. CD spectrum analysis showed that 32 kD protein had the features of globulin. The maximum ultraviolet absorbance was at 282 nm. Its GTPase activity was higher than its ATPase activity. It was activated by Ca~(2+) and Mg~(2+). The GTPase activity of the protein was insensitive to Na_(3)VO_(4) 100 #mu#mol/L and NEM 0.1 mmol/L, indicating that it was distinct from the motor proteins such as kinesin or dynamin.

Key words: Maize pollen;Low-molecular weight GTPase;Purification;Property

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引用本文：

王建华, 吴英杰, 吴显荣. 玉米花粉32 kD蛋白的纯化及性质[J]. 生理学报 1996; 48 (4): .

Wang Jianhua, Wu Yingjie, Wu Xianrong. Purification of 32 kD protein from maize pollen and study on some of its properties. Acta Physiol Sin 1996; 48 (4): (in Chinese with English abstract).