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OALib Journal期刊
ISSN: 2333-9721
费用：99美元
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嗜热子囊菌光孢变种β-葡萄糖苷酶的基因克隆、表达及酶学性质分析
DOI: 10.3724/SP.J.1145.2009.00549, PP. 549-553
Keywords: 嗜热子囊菌光孢变种,表达,纯化,毕赤酵母,β-葡萄糖苷酶
Abstract:
从嗜热子囊菌光孢变种(thermoascusaurantiacusvar.levisporus)rna中通过rt-pcr克隆出β-葡萄糖苷酶基因bglⅰ的全长序列，cdna序列为2672bp，genbank登录号为eu269025，将该片段插入巴斯德毕赤酵母pichiapastoris分泌型表达载体ppic9k中，获得重组质粒ppic9k/bgl，经线性化后用电穿孔法导入毕赤酵母gs115中，在醇氧化酶aox1基因启动子作用下，获得高效表达β-葡萄糖苷酶的毕赤酵母工程菌株.经deae-sepharosefastflow阴离子层析纯化了该重组表达蛋白.sds-page测得该重组蛋白相对分子质量(mr)约为120×103.经甲醇诱导，培养基中β-葡萄糖苷酶的活力可达1.2u/mg，小规模发酵量达0.45mg/ml.该酶的最适反应温度为60℃，最适反应ph为5.0.于70℃保温30min仍保持80%的酶活力，具有较高的热稳定性，在ph3.0~9.0的条件下酸碱耐受性强.图6表1参22
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