Bacillus flexusβ-淀粉酶低pH值突变体的构建及在麦芽糖制备中的应用

Construction of Lower Optimum pH Mutants ofBacillus flexusβ-Amylase and Its Application in Maltose Production

DOI：10.3969/j.issn.1673-1689.2019.10.015

中文关键词: β-淀粉酶 定点突变 麦芽糖 弯曲芽孢杆菌 酶转化

英文关键词: β-amylase,site-directed mutagenesis,maltose,B. flexus,enzyme conversion

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中文摘要:

工业上将β-淀粉酶与其他淀粉水解酶进行复配可获得高纯度的麦芽糖浆。相对于目前常用的植物来源β-淀粉酶，微生物β-淀粉酶具有生产工艺简单、不受原料限制、质量稳定、纯度高等优势，但其最适作用pH多为6.0~8.0，难以与其他淀粉水解酶（pH多为4.5~5.5）进行复配。本实验室前期获得的弯曲芽孢杆菌β-淀粉酶其最适pH值为7.0，本研究在此β-淀粉酶基础上构建T47K，Y164K，L396K三个突变体，其最适pH由突变前的7.0分别下降为6.0，4.5，5.5，其中L396K的最适温度也由突变前的50 ℃提高为60 ℃。以马铃薯淀粉（10% w/v）为底物进一步对L396K制备麦芽糖的转化条件进行探索与优化。结果表明，在反应初始pH为 5.5，温度60 ℃，β-淀粉酶加酶量为100 U/g干淀粉时麦芽糖最大转化率达到80.2%，符合高纯度麦芽糖浆的生产要求。

英文摘要:

Beta-amylase and other starch hydrolases were complexed to produce high purity maltose syrup in industry. Compared with plant β-amylase，the microbial β-amylase has the advantages of high purity，little limitation with the raw materials，and the suitability for large-scale production. However，the optimum pH of the microbial β-amylase is 6.0~8.0，which is difficult to complex with other starch hydrolases. In the pvious study，the optimum pH ofBacillus flexusβ-amylase was 7.0. In the present study，threeB.flexusβ-amylse mutants T47K，Y164K，L396K were constructed and the optimal pH were changed from 7.0 to 6.0，4.5 and 5.5 respectively. At the same time，the optimum temperature of L396K was increased from 50 ℃ to 60 ℃. The production conditions of maltose using L396K β-amylase was also optimized. When using potato starch（10% w/v） as the substrate under the condition of pH 5.5 and temperature 60 ℃，the yield of maltose reached 80.2%，which satisfied the production requirement of high purity maltose syrup.

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