大豆液泡膜V型H+ ATPase是ATPases中的一种 ,它在植物细胞的生长发育中有重要的作用 .利用竹红菌乙素 (HB)和KI这两种分别猝灭蛋白质疏水区域内源荧光和亲水区域内源荧光的荧光猝灭剂 ,在不同pH值、温度条件下对纯化的大豆液泡膜V型ATPase进行荧光猝灭实验 ,初步探讨了V型H+ ATPase的水解活性同其蛋白质折叠状态间的关系 .研究表明 ,通过比较不同 pH值、温度条件下蛋白质疏水区域和亲水区域内源荧光的荧光猝灭常数 (KSV) ,发现当环境pH值、温度偏离酶的最适 pH值和温度时 ,蛋白质的内源荧光强度降低且疏水区域和亲水区域内源荧光的荧光猝灭常数 (KSV)降低 ,说明伴随着酶的水解活性降低 ,蛋白质的折叠状态发生了变化 .我们认为蛋白质在膜内的折叠状态变化是酶失活机制的一个重要方面 ,为植物的抗冻和抗盐研究提供了一定的参考 Soybean tonoplast V-type H + ATPase is one of ATPases, which plays an important role in the growth and development of plant cells.Using both Hb and KI to quench endogenous fluorescence of protein hydrophobic region And fluorescence quenching agent of endogenous fluorescence in hydrophilic region, fluorescence quenching experiment was carried out on purified somatic V-ATPase of soybean so as to discuss the relationship between the hydrolytic activity of V-type H + ATPase and its protein folding The results show that by comparing the fluorescence quenching constants (KSV) of endogenous fluorescence in protein hydrophobic regions and hydrophilic regions at different pH values ​​and temperatures, it is found that when the environmental pH and temperature deviate from the optimal enzyme pH value And temperature, the endogenous fluorescence intensity of protein decreased and the fluorescence quenching constant (KSV) of endogenous fluorescence in hydrophobic and hydrophilic regions decreased, indicating that the protein folding state changed with the decrease of enzymatic hydrolysis activity. The change of the protein folding state in the membrane is an important aspect of the enzyme inactivation mechanism and provides a reference for the study of plant freezing and salt resistance