乙酰辅酶A含量对酿酒酵母乙酸乙酯合成的影响

通过缺失乙酰辅酶A水解酶（ACH）基因和过表达乙酰辅酶A合成酶（ACS）基因技术提高酿酒酵母合成乙酰辅酶A（acetyl- CoA）能力的同时，过表达醇酰基转移酶（ATF）基因，提高乙酸乙酯合成能力，并考察acetyl-CoA含量对酿酒酵母合成乙酸乙酯能力的影响。结果表明，敲除ACH1基因、且在敲除ACH1基因基础上过表达ACS1、ACS2基因均能提高酿酒酵母Acetyl-CoA含量，进而提高乙酸乙酯含量。较亲本菌株α5，缺失突变株α5ΔACH1、重组菌株α5-A1、α5-A2的Acetyl-CoA的含量均分别提高了52.5%、80.33%、52.79%，乙酸乙酯含量分别提高10.59%、26.12%、23.70%。在敲除ACH1基因、过表达ACS1和ACS2基因的基础上同时过表达ATF1基因，得到工程菌株A1-ATF1和A2-ATF1，较亲本菌株α5，乙酸乙酯含量分别提高226.09%、530.43%、289.57%，工程菌株A1-ATF1乙酸乙酯产量最高，为72.52 mg/L。研究表明，提高乙酰辅酶A含量能够促进乙酸乙酯的合成，为提高乙酸乙酯生成量提供了新思路。

Effect of acetyl-CoA content on ethyl acetate synthesis in Saccharomyces cerevisiae

To increase the synthesize ability of acety-CoA with Saccharomyces cerevisiae, the acety-CoA hydrolase (ACH) gene was deleted and the acetyl-CoA synthetase (ACS) gene was overexpressed. Meanwhile, overexpressing the alcohol acyltransferase (ATF) gene increased the ethyl acetate synthesize ability, and then the effect of acetyl-CoA content on the synthesis of ethyl acetate by S. cerevisiae was investigated. The results showed that both the gene ACH1 deleting and the genes ACS1/ACS2 overexpressing on the basis of deleting gene ACH could enhance the supply of acetyl-CoA for overproduction of ethyl acetate. The acety-CoA production of deleting mutant strain α5ΔACH1 and recombinant strains α5-A1and α5-A2 represented an increase of 52.5%, 80.33% and 52.79% compared with parental strain α5, respectively. Then the ethyl acetate production increased by 10.59%, 26.12% and 23.70%, respectively. Furthermore, engineered strains A1-ATF1 and A2-ATF1 were constructed with the deletion of gene ACH1, and simultaneously overexpressing of gene ACS1 and ACS2, the ethyl acetate production increased by 226.09%, 530.43% and 289.57% compared with the strain α5. The largest content of ethyl acetate was 72.52 mg/L by engineered strain A1-ATF1. The study showed that increasing the content of acetyl-CoA could improve the synthesis of ethyl acetate and it provided a new idea for increasing the production of ethyl acetate.