金黄色葡萄球菌低分子质量双特异性磷酸酶sL MWDSP的表达、纯化及性质  

Expression,Purification and Property of Low Molecular Weight Dual-specificity Phosphatase of Staphylococcus aureus

作　　者：郭伟伟[1] 倪晓华[1] 李继喜[2] 郑眉[2] 顾星[2] 季朝能[2] 

机构地区：[1]华东师范大学生命科学学院生物化学及分子生物学实验室,上海200062 [2]复旦大学生命科学学院遗传学研究所遗传工程国家重点实验室,上海200433

出　　处：《复旦学报（自然科学版）》2006年第3期293-297,共5页Journal of Fudan University：Natural Science

基　　金：上海市自然科学基金资助项目(04ZR14040)

摘　　要：双特异性磷酸酶是酪氨酸磷酸酶家族中的一员,它参与生物体内信号转导、生长调控、新陈代谢等许多基本的生理活动.金黄色葡萄球菌的低分子质量双特异性磷酸酶sLMWDSP(low molecular weight dual-specificityphosphatase,Staphylococcus aureus)基因从金黄色葡萄球菌cDNA库中克隆,并在大肠杆菌中表达.高纯度的sLMWDSP用亲和层析的方法纯化得到.酶学性质研究表明:sLMWDSP催化对硝基苯磷酸(-pnitrophenyl phos-phate,pNPP)水解反应的最适pH值为6.7,最适温度为35℃,且该酶的活性不依赖于金属离子.磷酸酶通用抑制剂岗田酸(Okadaic acid)、EDTA对sLMWDSP几乎没有抑制作用,但钒酸钠能明显地抑制该酶的活性.sLMWDSP对pSer/Thr以及pTyr的寡肽均有去磷酸化作用,这说明sLMWDSP是一个新的双特异性磷酸酶.Dual-specificity phosphatase is a member of protein tyrosine pbosphatase family and plays a key role on a series of fundamental biological functions in many organisms, including signal transduction, growth control, metabolism. sLMWDSP gene （low molecular weight dual-specificity phosphatase, Staphylococcus aureus） is cloned from Staphylococcus aureus and expressed in E. coli Rossetta and sLMWDSP is purified by affinity chromatography. The analysis of enzyme property shows that its optimum pH is 6.7, and the optimum temperature is 35 ℃ for the hydrolysis of pNPP （ pnitrophenyl phosphate）and the activity can not be enhanced by metal ions. sLMWDSP can not be inhibited by Okadaic acid and EDTA, but obviously can be inhibited by vanadate sodium, sLMWDSP shows phosphatase activity towards oligopetptides containing pSer/Thr and pTyr, indicating that sLMWDSP is a typical protein phosphatase with dual substrate specificity.

关 键 词：双特异性磷酸酶 对硝基苯磷酸 金黄色葡萄球菌 磷酸酶活性 

分 类 号：Q814[生物学—生物工程]