短短芽孢杆菌几丁质酶的分离纯化及酶学性质

Purification and Characterization of Chitinase from the Fermentation Broth of Brevibacillus brevis FM4B

探讨来源于短短芽孢杆菌FM4B的几丁质酶(chitinase)分离纯化过程及其性质。菌株FM4B摇瓶发酵液经过离心、乙醇分级沉淀及葡聚糖凝胶G-100层析纯化,用变性聚丙烯酰胺凝胶电泳确定其分子质量。结果获得几丁质酶纯品,酶的纯化倍数为7.19,酶活回收率为30.6%,分子质量为66 kD;酶活力在pH 5.0~9.0和80℃以下稳定,最适pH值为6.0,最适温度为50℃;Cu^(2+)、Hg^(2+)、pb^(2+)、Co^(2+)以及Zn^(2+)对该酶有强烈的抑制作用,Mg^(2+)和Ca^(2+)对该酶的活力具有一定的促进作用;该几丁质酶对多种霉菌有显著的抑菌效果。菌株FM4B分泌的几丁质酶稳定性好,且对多种霉菌抑制效果明显,具有较高的潜在利用价值。

In the present study,the purification and characterization of a chitinase from the fermentation broth of Brevibacillus brevis FM4 B was investigated by centrifugation,ethanol precipitation and Sephadex G-100 chromatography.Sodium dodecyl sulfate polyacrylamide gel electrophoresis（SDS-PAGE） was used to determine its molecular weight.Purified chitinase with a purification factor of 7.19 and a recovery rate of 30.6%was obtained.The molecular weight of the purified chitinase was approximately 66 kD.It had an optimum reaction temperature and pH of 50 ℃ and 6.0,respectively.The enzyme also showed good stability in the pH range of 5.0-9.0.Its activity could be inhibited by Cu2＋,Hg2＋,Pb2＋,Co2＋ and Zn2＋,but slightly activated by Mg2＋ and Ca2＋.In addition,the chitinase had obvious inhibitory effect on different fungi.The chitinase,possessing high thermal stability,good pH adaptability and significant antifungal effect,has great potential for practical applications.