温度和底物对大豆液泡膜H^+-ATPase二级结构的影响

TEMPERATURE AND SUBSTRATE ON THE EFFECTS OF CIRCULARDICHROISM OF TONOPLAST H^+-ATPase FROM SOYBEAN

利用圆二色性光谱 ,检测了纯化的大豆液泡膜H+ -ATPase在不同条件下蛋白二级结构的变化。液泡膜H + -ATPase的圆二色光谱对温度敏感 ,25°C、37°C分别保温10分钟和20分钟 ,208nm和222nm双负峰变小 ,酶蛋白α -螺旋含量减少 ,与25°C相比较 ,37°C保温时酶蛋白构象的变化更为剧烈、迅速。不同磷酸数目的腺苷酸处理 ,液泡膜H+ -ATPase的α -螺旋含量均降低 ,降低程度为ADP>AMP>ATP ,酶蛋白二级结构对底物磷酸基数目敏感 ;用含不同碱基的三磷酸核苷酸处理 ,ATPase的α -螺旋含量均降低 ,降低程度为GTP>CTP>ATP ,酶蛋白二级结构对底物碱基类型敏感。与最适底物ATP相比 ,其它底物对酶蛋白二级结构的影响更为剧烈。

The circular dichroism of purified tonopast H+-ATPase from soybean was studied under different conditions. Compared with the control, the double negative peaks at 208nm and 222nm of the CD spectra of the enzyme under 25°C and 37°C for 10 and 20 minutes declined, which meant the decreasing of the α-helix content. Twenty minutes later, CD of the enzyme became stable, in the following order: When the enzyme was treated with ADP、 AMP and ATP respectively, of ADP>AMP>ATP. This suggested the secondary structure of the enzyme was sensitive to the number of the phosphates of the nucleotide. The α-helix content also decreased when the enzyme was dealt with different nucleotide triphosphates respectively, in the order of GTP>CTP>ATP, which meant the conformation of the enzyme was sensitive to the bases of the nucleotides. ATP induced change of the conformation of the enzyme was smaller than the other substrates.