【摘要】 构建了蛇肌cDNA文库 ,用抗体筛库 ,克隆了肌酸激酶的cDNA ,测定了其核苷酸序列 ,并将完整的cDNA克隆到 pET11表达质粒 ,在大肠杆菌中获得高效表达。纯化的重组肌酸激酶 ,与组织酶的动力学性质表现出高度的一致性。同时 ,比较了蛇肌酸激酶与其他种属M型肌酸激酶的同源性 ,确定了在爬行动物中肌酸激酶存在M型。更多还原

【Abstract】 Snake muscle cDNA library was constructed, and an allelic cDNA codin g for M-type creatine kinase was cloned through screening library with antibody . A complete open reading frame(1 140 bp) codes for 380 amino acids which shows h igh homology with known creatine kinase isoenzyme. After cloning into expression vector pET11a, the snake muscle creatine kinase(SM-CK) was over-expressed in Es cherchia coli. Kinetical studies showed that purified recombinant SM-CK exhib ite d behavior similar to the tissue form. Evolutionary tree of M-type creatine kin a se was then constructed, and it was determined that this allelic cDNA of snake m uscle creatine kinase belonged to M-type situated between in mammals and birds. These findings, along with the following SM-CK gene regulation studies, may she d light on the role of creatine kinase in energy generation during hibernation as well as the molecular basis of myogenesis during ontogeny.更多还原