【摘要】 利用平板快速筛选法 ,从鱼腹中筛选到一株产生硫酸软骨素酶的细菌YH31 1 ,通过生物特性和生化反应试验考察 ,初步鉴定为温和气单胞菌 (Aeromonassobria)。培养至 36h时 ,该菌株产酶达到高峰期 ,培养液的酶活力达0 9U mL。利用超声波破碎菌体细胞 ,分别测定培养液和菌体细胞的酶活力 ,发现培养液的酶活力要远远大于菌体细胞的酶活力 ,显示该酶为分泌型表达 ,属于胞外酶。发酵液经离心后 ,上清液用硫酸铵分级沉淀 ,再分别经过CM Cellulose、QAE sephadexA5 0和SephadexG 1 5 0层析柱进行逐级分纯 ,并跟踪酶活 ,最后获得硫酸软骨素酶。SDS PAGE检测为单一条带 ,其分子量约为 80kD。更多还原

【Abstract】 StrainYH311, with a high level of chondroitinase productivity,was obtained from the intestinal contents of the fish by rapid plate screening. It was identified to be Aeromonas sobria according to the biological characteristics and biochemical reaction tests. It produced highest amount of enzyme at 36 hours′ incubation, which reached to 0 9U/mL. By comparing the extracellular and intracellular chondroitinase activity of the strain disrupted by sonication, it was found the former was much higher than the latter, which showed that YH311 secretorily expressed the chondroitinase and the chondroitinase was extracellular enzyme. Chondroitinase was prepared from the culture fluid by fractional precipitation with ammonium sulfate, chromatography on CM cellulose and QAE sephadex A50, and finally gel filtration on Sephadex G 150. SDS PAGE of the preparation gave a single band which show its molecular weight was about 80kD.更多还原