弱化乙醇途径关键酶活性减少酿酒酵母在丙酮酸发酵过程中副产物乙醇的积累

摘要

Four-carbon dicarboxylic acids are widely used in food,pharmaceutical,and chemical industries.Saccharomyces cerevisiae is regarded as the best candidate microorganism for producing four-carbon dicarboxylic acids,however,there is strong tendency for ethanol formation under fully aerobic conditions when sugar are present in excess in batch culture,which leads to the loss of carbon flow and cofactor when the desired product are four-carbon dicarboxylic acids.In this study,it showed that deletion of both the pyruvate decarboxylase 1 (PDC1) and the alcohol dehydrogenase 1 (ADH1) genes significantly reduced cell growth.Furthermore,deletion of both PDC1 and ADH1 also reduced Pdc and Adh enzyme activities,decreased ethanol production,and increased pyruvate production under optimal fermentation conditions,which led to 63.8％ decrease of ethanol and 2.0 g/L pyruvate accumulation compared to the control.Therefore,the strategy might be valuable to reduce carbon flow toward ethanol to improve production of C4-dicarboxylic acids and other organic acids.%C4二元羧酸广泛应用于食品、医药和化工等产业.酿酒酵母被认为是生产C4二元羧酸的潜在最适微生物,但在高糖浓度及有氧条件下,它会产生大量的乙醇,对于以C4二元羧酸为目标产物来说,这会导致碳代谢流和辅助因子的损失.通过敲除PDC1和ADH1两个关键基因可同时弱化Pdc和Adh的活性,但导致菌体生长较弱.在最佳的培养基组成和培养条件下,双基因突变株的最大生物量与野生型菌株接近,发酵48 h时,最大乙醇产量为2.1 g/L,较野生型菌株最大乙醇产量下降了63.8％,此时,丙酮酸产量为2.0 g/L,而野生型菌株几乎不产丙酮酸.通过弱化乙醇途径关键酶活性可以有效减少乙醇形成,为解决利用酿酒酵母生产C4二元羧酸及其它有机酸中副产物问题提供了一个可供选择的策略.