泥蚶(Tegillarca granosa)精氨酸激酶的基因克隆与表达  

CLONING AND OVEREXPRESSION OF TEGILLARCA GRANOSA ARGININE KINASE

作　　者：李松伟[1] 李晔[1] 李成华[1] 李太武[1,2] 苏秀榕[1] 

机构地区：[1]宁波大学生命科学与生物工程学院,宁波315211 [2]宁波城市职业技术学院,宁波315100

出　　处：《海洋与湖沼》2011年第1期119-123,共5页Oceanologia Et Limnologia Sinica

基　　金：国家自然科学基金资助项目;30771665号;长江学者和创新团队发展计划资助项目;2007-2009;浙江省科技厅重点科研社会发展项目;2005C23083号

摘　　要：利用PCR扩增技术,从泥蚶的cDNA文库中克隆出了精氨酸激酶(arginine kinase)基因,并进行了原核表达。精氨酸激酶是调节能量代谢的重要酶类,在无脊椎动物体内能量平衡过程中起重要作用。结果表明,泥蚶的精氨酸激酶全长1601bp,包括5′非翻译区(5′-UTR)序列332bp,3′非翻译区(3′-UTR)序列246bp、1023bp的开放阅读框序列(ORF),编码340个氨基酸,具有精氨酸激酶典型的酶活性部位氨基酸序列(CPTNLGT)。构建了原核表达质粒AK-pET-28a(+)转化表达菌株,经IPTG诱导表达后,获得了与预期的分子量大小一致的表达产物。利用该目的蛋白制备了泥蚶精氨酸激酶的特异性多克隆抗体,经Western blot证明该蛋白为精氨酸激酶。In vertebrate, arginine kinase is a phosphotransferase that plays a critical role in maintaining equilibrium in energy metabolism. We isolated full-length cDNA of arginine kinase from Tegillarca granosa. It is 1601 bp long, including 203 bp of 5＇ untranslated region （5＇-UTR） sequence and 438 bp of 3＇ untranslated region （3＇-UTR） sequence. It encodes 319 amino acids with arginine kinase activity in part of the amino acid sequence （CPTNLGT）. And the resulting product was then cloned into the expression vector. The prokaryotic expression plasmid AK-Pet-28a（＋） containing the intact ORF was constructed and the target protein was induced with IPTG. The overexpressed protein was with the expected size. This study provided information further built on the foundation in understanding the function of arginine kinase in energy metabolism and host defense.

关 键 词：泥蚶 精氨酸激酶 基因克隆与表达 多克隆抗体 

分 类 号：Q75[生物学—分子生物学]