Zhang Feng, Wang Zhaoyue, Liu Nan, He Xiuping, Zhang Borun
Laboratory of Molecular Genetics and Breeding of Yeast, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
Sheng Wu Gong Cheng Xue Bao. 2008 May;24(5):837-43.
Saccharomycopsis fibuligera possesses high alpha-amylase and glucoamylase activities that enable it to utilize raw starch as a carbon source. A expression cassette containing the promoter sequence of 3-phosphogylycerate kinase gene (PGK1p), the alpha factor signal sequence from Saccharomyces cerevisiae and the alpha-amylase coding sequence of S. fibuligera was constructed. The alpha-amylase expression cassette was inserted in the ILV2 locus of industrial brewer's yeast strain YSF-5 encoding alpha-acetolactate synthase (AHAS) by homologous recombination. The transformed yeast strain was selected on the media with starch as the sole carbon source and verified by PCR. The transformant exhibited secretive alpha-amylase activity, low AHAS activity and reduced diacetyl production. Effects of temperature, pH, and metal ions on the activity of the alpha-amylase expressed by the transformant were examined. The fermentation performance of host strain YSF-5 and the transformant was also examined.
扣囊复膜孢酵母具有较高的α-淀粉酶和糖化酶活性，使其能够利用生淀粉作为碳源。构建了一个表达盒，其包含3-磷酸甘油酸激酶基因（PGK1p）的启动子序列、来自酿酒酵母的α因子信号序列以及扣囊复膜孢酵母的α-淀粉酶编码序列。通过同源重组将α-淀粉酶表达盒插入工业酿酒酵母菌株YSF-5编码α-乙酰乳酸合酶（AHAS）的ILV2基因座中。在以淀粉为唯一碳源的培养基上筛选转化后的酵母菌株，并通过PCR进行验证。该转化体表现出分泌性α-淀粉酶活性、低AHAS活性和降低的双乙酰产量。研究了温度、pH和金属离子对该转化体表达的α-淀粉酶活性的影响。还研究了宿主菌株YSF-5和转化体的发酵性能。
