摘要
解木聚糖类芽孢杆菌(Paenibacillus xylanilyticus)发酵液经硫酸铵分级沉淀、HiPrep26/10 Desalting柱脱盐、HiPrepDEAE FF16/10阴离子交换柱、HiPrep 16/60 Sephacryl S-100凝胶柱、HiPrep 16/10 Source 30S阳离子交换柱等,最终纯化出单一组分的木葡聚糖酶,经过SDS-PAGE电泳分析,此木葡聚糖酶相对分子量约为39 kD。该菌所产木葡聚糖酶的最适反应温度是50℃,在60℃以下较稳定;最适反应pH是7.0,在pH5.0-10.0范围内酶活力较为稳定。酶的动力学研究显示Km为65 g/L,Vmax为6.49μmol/min,kcat=10.86 s-1。底物特异性研究表明对木葡聚糖具有较高比活力。酶蛋白经质谱分析,比对结果显示与来源于Paenibacillus pabuli的木葡聚糖酶有较高同源性。本研究为首次报道解木聚糖类芽孢杆菌(P.xylanilyticus)产木葡聚糖酶。
Xyloglucanase had been extracted from the culture medium fermented by Paenibacillus xylanilyticus. Isolated and purified by methods of ammonium sulfate precipitation, HiPrep26/10 Desahing gel layer chromatography, HiPrepDEAE FF16/10 anion exchange column, HiPrep 16/60 Sephaeryl S-100 gel layer chromatography and I-IiPrep 16/10 Source 30S cation exchange column. The molecular weight of xyloglucanase was about 39 kD, identified by SDS-PAGE. The optimal reaction temperature of xyloglucanase was 50℃, and there was good stabi-lity under 60℃. The optimal reaction pH of xyloglucanase was 7.0, and there was good stability between pH5.0 and pH10.0. The dynamic study demonstrated that the Km value of xyloglueanase was 65 g/L, the maximum reaction velocity （ Vmax ） was 6.49 μmol/min, and the kcat, was 10.86 s^-1 . The substrate specificity of the xyloglueanase indicated that the xylogluean had much higher specific activity than other substrates. The purified enzyme was analyzed by mass spectrometry, and the result showed that the purified enzyme had high homology with xyloglucanase from Paenibacillus pabuli. It was the first report about xyloglucanase product from PaenibaciUus xylanilyticus.
出处
《生物技术通报》
CAS
CSCD
北大核心
2012年第7期126-133,共8页
Biotechnology Bulletin
基金
国家"863"计划课题资助项目(2011AA100905)
关键词
解木聚糖类芽孢杆菌
木葡聚糖酶
分离纯化
酶学性质
质谱分析
Paenibacillus xylanilyticu
Xyloglucanase Purification Enzyme characterieation Mass spectrometry
