变幻青霉氨基甲酸乙酯降解酶产酶条件优化及酶的底物特异性

Optimization of Fermentation Conditions of P.variabile JN-A525 and Substrate Specificity of Urethanase

DOI：10.3969/j.issn.1673-1689.2013.06.008

中文关键词: 氨基甲酸乙酯降解酶 优化 纯化 底物特异性

英文关键词: urethanase optimization purification substrate specificity

基金项目:国家973计划项目(2012CB720802)

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中文摘要:

在单因素优化基础上,运用正交试验设计理论,优化氨基甲酸乙酯降解酶产生菌P.variabile JN-A525的产酶条件。优化发酵培养基配方为:葡萄糖20 g/L,牛肉膏10 g/L,KH2PO45g/L、NaCl 1 g/L、KCl 5 g/L;发酵条件为:初始pH 6.0、培养温度32℃、接种体积分数3%、250 mL摇瓶装量70 mL、摇床转速100 r/min,在此优化条件下酶活比优化前提高了3.47倍。对经超声破碎、(Na)2SO4盐析、Superdex G-75凝胶过滤层析步骤初步纯化的酶之底物特异性的研究表明,在模拟黄酒体系下,该酶对尿素和氨基甲酸乙酯有相对强的降解作用,对一些酯类和氨基酸没有降解作用。

英文摘要:

The fermentation process for P.variabile JN-A525 was optimized by single factor experiment and orthogonal tests.The results showed that the medium was composed of glucose 20 g/L,beef extract 10 g/L,KH2 PO4 5 g/L,NaCl 1 g/L and KCl 5 g/L.The optimum fermentation conditions was 32 ℃,inoculum volume 3%,the optimum amount in flask 70/250 mL and the rotation speed of 100 r/min.Under these optimal conditions,the highest urethanase activity is 3.47 fold more than that of previous cinditions.After sonication methods,sodium-sulfate salting-out and superdex G-75 gel filtration chromatography,the urethanase from Penicillium variabile JN-A525 is further determined substrate specificity.Under simulated conditions,the urethanase has a relatively strong catalytic activity to urea and EC.Meanwhile,urethanase cannot catalyze the decomposition of amino acids and esters.

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