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中文摘要：
【目的】进一步提高光滑球拟酵母（Torulopsis glabrata）葡萄糖代谢速率及丙酮酸生产强度。【方法】将源于荚膜胞浆菌（Histoplasma capsulatum）的编码选择性氧化酶的AOX1基因过量表达于T.glabrata中,获得了一株线粒体内NADH氧化途径发生改变且胞内总NADH氧化酶活性提高1.8倍的重组菌株AOX。【结果】与出发菌株CON比较,细胞浓度以及发酵周期降低了20.3%和10.7%,而平均比葡萄糖消耗速率和丙酮酸合成速率分别提高了34.7%和54.1%。其原因在于NADH选择性氧化酶过量表达,导致胞内NADH/NAD＋比率和ATP含量分别降低了74.7%和52.9%,而磷酸果糖激酶和丙酮酸激酶活性分别提高了185.0%和28.1%。【结论】构建线粒体内非产能的NADH氧化途径,加速NADH氧化的同时降低ATP合成,能有效地提高酵母细胞糖酵解代谢速度及目标代谢产物的生产强度。
英文摘要：
[Objective] This study aimed at increasing the glycolytic flux and pyruvate productivity in Torulopsis glabrata with glucose as carbon source. [ Methods ] For this target, we introduced a mitochondrial located alternative oxidase encoded by Histoplasma capsulatum AOX1 gene into T. glabrata, and a mutant strain named as AOX with total NADH oxidase activity was 1.8-fold higher than that of the parent stain, was achieved. [ Results] The heterologous expression of NADH alternative oxidase resulted in decrease of the dry cell weight and fermentation time by 20.3% and 10.7%, but the specific rate of glucose consumption and pyruvate production increased 34.7 % and 54.1% higher, respectively. The reasons for high glycolytic flux were the intracellular NADH/NAD ＋ ratio and ATP concentration decreased 74.7 % and 52.9 % respectively, the specific activity of phosphofructokinase, pyruvate kinase increased 185.0% and 28.1%. [ Conclusion]Introduction of a novel NADH oxidation pathway by alternative oxidase can efficiently increase the rate of glucose consumption and the target metabolite productivity.
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