淀粉液化芽孢杆菌BS5582产蛋白酶的酶谱分析与鉴定
Zymogram analysis and identification of the proteases from Bacillus amyloliquefaciens BS5582
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摘要: 本研究对淀粉液化芽孢杆菌BS5582分泌的蛋白酶进行快速分析与鉴定。测定菌株胞外蛋白酶酶活,结果显示蛋白酶在中性条件下活力最高,且在发酵过程中出现三个酶活高峰,表明菌株可能分泌多种蛋白酶。蛋白酶酶谱分析发现共有五条蛋白水解条带。通过质谱鉴定这些水解条带对应非变性-SDS-PAGE凝胶上的蛋白条带,表明该菌株可能产Bacillopeptidase F、胞外中性金属蛋白酶和枯草杆菌蛋白酶Bpn’这三种蛋白酶,其中Bacillopeptidase F和胞外中性金属蛋白酶存在两种形式。结果显示酶谱分析法结合质谱鉴定能快速鉴定菌株BS5582分泌的胞外蛋白酶的多种复杂形式,是一种快速鉴定多种蛋白酶的有效方法。Abstract: The proteases secreted by strain BS5582 were rapidly identified by zymogram method in this study.The highest extracellular protease activity was detected under neutral condition. The three peaks of protease during fermentation process indicated that BS5582 might secrete several proteases.The zymography displayed five clear bands of casein hydrolysis on the gel. Protein bands on the non- denaturing- SDS- PAGE gel corresponding to those on the zymography gel were identified by mass spectroscopy. The results showed that strain BS5582 secreted three kinds of proteases.They were Bacillopeptidase F ( Bpr) , extracellular neutral metalloprotease ( Mpr) and Subtilisin Bpn '. Bpr and Mpr had two forms in the medium, respectively. The different forms of extracellular proteases secreted by BS5582 could be rapidly identified by the zymography method combined with application of mass spectrometry.Therefore, it was an efficient and rapid method to identify mixture of proteases.
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