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目的:纯化灭蚊真菌贵阳腐霉胞外蛋白酶类枯草杆菌蛋白酶,并测定其分子量。方法:培养贵阳腐霉菌丝体,用几丁质诱导,使其产生大量胞外酶,对其产酶条件进行了部分优化。将粗酶液经过脱盐,浓缩进一步纯化,再过Sephadex G-100柱子层析并进行SDS-PAGE,分离类枯草杆菌蛋白酶。结果:(1)以几丁质为唯一碳氮源,在26℃,初始pH值6.0,1%的菌丝接种量为最佳诱导条件;(2)分离得到的类枯草杆菌蛋白酶,并测定其分子量约为33kDa。结论:类枯草杆菌蛋白酶纯化成功,结果较理想,为进一步研究提供依据。
Objective: To purify the protease subtilisin from Pythium guiyangense, a mosquito-killing fungus, and determine its molecular weight. Methods: Pythium guiyan mycelium was cultured and induced by chitin to produce a large number of extracellular enzymes, and the conditions of its production were partially optimized. The crude enzyme solution was desalted, concentrated for further purification, and then subjected to Sephadex G-100 column chromatography and subjected to SDS-PAGE to separate the subtilisin-like protease. Results: (1) The optimum conditions for the inoculation of mycelium with chitin as sole carbon and nitrogen source at 26 ℃ and initial pH value of 6.0 and 1% were obtained. (2) Subtilisin-like protease was isolated and assayed Its molecular weight is about 33 kDa. Conclusion: The subtilisin-like protease has been successfully purified and the result is satisfactory. It provides the basis for further study.