来源于假单孢菌206植酸酶基因的克隆、表达及酶学性质研究  

Gene Cloning,Expression and Characterization of an HAP Phytase from Pseudomonas fluorescens 206

作　　者：李雅楠[1] 黄火清[2] 姚斌[2] 赵青[1] 刘昆[1] 马文康[1] 

机构地区：[1]广州医学院生物化学与分子生物学教研室,广州510182 [2]中国农业科学院饲料研究所,北京100081

出　　处：《生物技术通报》2013年第8期139-144,共6页Biotechnology Bulletin

基　　金：广州医学院科研项目(2006GD058)

摘　　要：从浙江嘉兴人工鱼塘底泥样品筛选到产植酸酶活性较高的假单孢菌(Pseudomonas fluorescens)206。通过简并PCR和TAIL-PCR技术从Pseudomonas fluorescens206菌株基因组DNA中克隆得到一个新的编码组氨酸酸性植酸酶(HAP)的基因,命名为phyP。该基因ORF全长1 284 bp,编码了427个氨基酸和一个终止密码子,前24个氨基酸为信号肽。将phyP在大肠杆菌中表达,重组蛋白经硫酸铵分级沉淀和阴离子交换柱纯化后达到电泳纯。对其酶学性质分析表明:重组植酸酶PhyP最适pH为5.5,在pH3.5-7.5的条件下具有较好的稳定性;重组植酸酶PhyP最适温度为45℃,在25℃时也具有较高的酶活性,因此植酸酶PhyP在饲料行业,尤其是水产行业具有潜在的应用前景。A Pseudomonas fluorescens 206 strain producing HAP phytase was isolated from the bottom mud of artificial fish ponds in Jiaxin city.A novel HAP phytase encoding gene phyP was cloned by degenerate PCR and TAIL-PCR.The full length gene consist 1 284 bp and encodes 427 amino acids,including a putative signal peptide of 24 residues.The gene phyP was expressed in E.coli.The recombinant enzyme PhyP was purified to homogeneity by ammonium sulfalte precipitation and anion exchange chromatography.The optimal pH of recombinant PhyP was pH5.5,and the enzyme showed high stability at pH3.5-7.5.The optimal temperature of PhyP was found to be 45℃,and the enzyme had a higher activity at 25℃.These superior properties of PhyP make it advantageous for applying in feed industries,especially fisher industry.

关 键 词：假单孢菌(Pseudomonas fluorescens) 植酸酶 基因克隆与表达 酶学性质 

分 类 号：S816.3[农业科学—饲料科学] S963[农业科学—畜牧学]