木霉菌发酵羊胃容物产木聚糖酶的分离纯化及特性

Purification and Characterization of Xylanase Produced by Trichoderma in Solid-state Fermentation of Sheep Gastric Contents

目的：为城市屠宰场牛羊胃容物类的生物垃圾型可再生资源的潜在工业化开发应用（城市无害鲜生物垃圾变废为宝的低碳生物循环经济工程）提供了部分可靠科学依据。方法：M-112木霉菌固态发酵羊胃容物的粗酶提取液，经20-60％饱和度的硫酸铵粗分级分离、SephadexG-25脱盐、SephadexG-150到SephadexG-100分子筛顺序纯化，通过蛋白质测定、酶活测定、SDS—PAGE和底物-PAGE电泳等。结果：制备到一种分子量约为64．7kDa的电泳纯木聚糖酶，其回收率为5．59％，纯化倍数10．43。比活力为253．36IU／mg。纯化出的木聚糖酶，经性质研究表明，其Km=11．20g／L，Vmax=0．70μmol／min；最适温度为65℃，最适pH为6．O，该酶在中温条件下稳定性好，在酸性至弱碱性（pH3．0～8．0）条件下稳定性好。结论：采用改良的凝胶色谱技术，首次纯化出木霉菌固态发酵羊胃容物所产的木聚糖酶。

Objective ：The date of this study provided some reliable scientific basis for potential industrial development application of biological waste - based renewable resources. Method：The xylanase was separated and purified from a culture filtrate of Tr/choderma M - 112 in solid - state fermentation of sheep gastric contents through ammonium sulfate precipitation,Sephadex G - 25 column chromatography and Sephadex G - 150→Sephadex G - 100 successive column chromatography. The purified xylanase was proved by SDS - PAGE and Native PAGE to be a single composition. Result：The xylanase＇ s molecular mass was 67.4kDa. The recovery rate was 5.59%. Purification fold was 10.43 and the specific activity was 253.36IU/mg. By study of characterization,Its Km is 11.20g/L,and Vmax was 0. 70μmol/min, Optimum temperature was 65℃, the optimum pH was 6.0 ;This xylanase had good stability at medium temperature. It also had good stabil- ity at pH 3.0～8.0. Conclusion： The xylanase produced by Trichoderma in solid - state fermentation of sheep gastric contents was first time purificated using improved gel chromatography technology.