摘要
目的通过比较HSP70野生型和突变型蛋白的自身磷酸化及ATP分解合成反应的不同,探讨HSP70的作用机制。方法运用基因定位诱导突变技术,将89和227位组氨酸分别替代以丝氨酸(H89S,H227S),分析野生型和突变型HSP70蛋白的自身磷酸化过程,对两种蛋白的ATP分解和合成变化进行研究。结果突变的H89S蛋白自身磷酸化、ATP分解和合成受到抑制,有剂量反应关系存在;野生型和H227S蛋白的ATP交换过程未受到影响。结论89位组氨酸点突变能显著降低ATP酶交换反应,但它的自身磷酸化可能并非必须的介导位点或只是一个选择性的功能侧链。
Objective To evaluate autophosphorylation, ATPase and ATP synthesis of wild type and mutant HSP70. Methods Select two most possible sites of histidine, as intermediate phosphorylation sites, i.e. His-89 and His-227, based on the hypothesis of the transfer of γ-phosphoryl groups and replaced by serine site-directed mutagenesis. Results An acid labile autophosphorylation intermediate of HSP70 and its CDP-dependent dephosphorylation were detected in wild type HSP70, which were markedly suppressed in mutation on H89S of HSP70, but not H227S mutation. The ATPase activity and ATP synthesis activity of HSP70 were almost completely suppressed in mutation on H89S. Conclusion Point mutations of His-89 significantly lower ATPase activity. Mutational studies suggest that phosphor-histidine may not be an obligatory intermediate, or an alternative sideehain to serve as a phosphate aeeeptor.
出处
《上海第二医科大学学报》
CSCD
北大核心
2005年第9期925-927,共3页
Acta Universitatis Medicinalis Secondae Shanghai
