CAJ | 학술논문

从全国各地采集豆豉样品,经富集培养并利用纤维蛋白平板法获得一株形态与现存产纤溶酶微生物差异较大的菌株HS9。通过传统方法、化学方法以及16SrRNA序列分析对HS9进行分类鉴定,属于Pseudomonas aeruqinosa,是未见报道的产豆豉纤溶酶菌株。发酵培养HS9获得粗酶,经20%~70%硫酸铵梯度盐析、SephadexG-75凝胶过滤以及CM-Sepharose Fast Flow阳离子交换层析分离纯化后,得到了电泳纯酶。通过SDS-PAGE了解该酶分子量约为34kD,pH8.0~8.5时酶活性最高,最适作用温度48℃,作用方式为直接水解纤维蛋白,胃蛋白酶抑制剂在工作浓度1μmol/L时能完全抑制其活性,推测该酶为天冬氨酸蛋白酶,是一种新型的豆豉纤溶酶。
종전국각지채집두시양품,경부집배양병이용섬유단백평판법획득일주형태여현존산섬용매미생물차이교대적균주HS9。통과전통방법、화학방법이급16SrRNA서렬분석대HS9진행분류감정,속우Pseudomonas aeruqinosa,시미견보도적산두시섬용매균주。발효배양HS9획득조매,경20%~70%류산안제도염석、SephadexG-75응효과려이급CM-Sepharose Fast Flow양리자교환층석분리순화후,득도료전영순매。통과SDS-PAGE료해해매분자량약위34kD,pH8.0~8.5시매활성최고,최괄작용온도48℃,작용방식위직접수해섬유단백,위단백매억제제재공작농도1μmol/L시능완전억제기활성,추측해매위천동안산단백매,시일충신형적두시섬용매。
Douchi, a kind of Chinese traditional fermented soybean food, were collected from China, and a new Douchi fibrinolytic enzyme-producing strain named HS9 was isolated from them through enrichment culture and fibrin plate. It identified as Pseudomonas aeruqinosa by classical classification system, chemical approaches and 16S rDNA sequencing. Crude enzyme, obtained from HS9 broth supernatant, was purified by 20%~70% ammonium sulfate fractional precipitation, Sephadex G-75 profile and CM-Sepharose Fast Flow profile. SDS-PAGE revealed the molecular weight of this enzyme was 34 kD. The optimal pH value was pH 8.0~8.5 and the optimal temperature was 48℃, respectively. The mode of fibrinolytic action is lysis fibrin directly. Pepstatin inhibited its activity completely at 1 μmol/L. The enzyme may be an aspartic acid enzyme and different from existent Douchi fibrinolytic enzyme.