缺失氨肽酶N的变铅青链霉菌的构建和鉴定  

CONSTRUCTION AND CHARACTERIZATION OF AMINOPEPTIDASE N DEFICIENT STREPTOMYCES LIVIDANS TK24

作　　者：洪斌[1] 李元[1] Jozef Anné 

机构地区：[1]中国医学科学院医药生物技术研究所,北京100050 [2]LaboratoryofBacteriology,RegaInstitute

出　　处：《微生物学通报》2003年第1期14-19,共6页Microbiology China

基　　金：国家自然科学基金资助项目 (No 30 0 70 0 0 9);中国与比利时弗莱芒区国际科技合作项目资助 (No BIL97/ 4 2 )~~

摘　　要：用PCR方法扩增变铅青链霉菌 (Streptomyceslividans)TK2 4氨肽酶N基因 ,体外插入卡那霉素抗性基因进行失活 ,然后利用不含链霉菌复制起点的重组质粒 pPEPN KAN进行同源重组 ,获得了氨肽酶N缺失的菌株PEPN- 。突变株的胞外氨肽酶N活性与原株基本相同 ,而胞内氨肽酶N的活性明显低于原株 ,为原株的 4 2 5± 5 7%。N terminal degradation has been noted for some homologous and heterologous proteins expressed in Streptomyces lividans , and genes of some aminopeptidases have been cloned Pep N is the one which has broad substrate specificities and the major leucine aminopeptidase activity in S lividans We cloned the pepN gene by PCR and inactivated it by homologous recombination to replace the wild type chromosomal pepN with a mutant gene in which kanamycin resistant gene was inserted The recombinational exchange of mutant and wild type allele was effected using a non replicating plasmid pPEPN KAN (derivative of pUC19) The intracellular and extracellular PepN activities of wild type and negative strains were detected The ability of the negative strains (designated PEPN -) to hydrolyze Leu pNA was significantly reduced compared to the wild type

关 键 词：变铅青链霉菌 氨肽酶N 缺失菌株 

分 类 号：Q939.9[生物学—微生物学]