Repression of mitochondrial metabolism for cytosolic pyruvate-derived chemical production in Saccharomyces cerevisiae
Repression of mitochondrial metabolism for cytosolic pyruvate-derived chemical production in Saccharomyces cerevisiae
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DOI：
10.1186/s12934-019-1226-6
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发表时间：
2019-10-15
影响因子：
6.4
通讯作者：
Shimizu, Hiroshi
中科院分区：
文献类型：
--
作者：
Morita, Keisuke;Matsuda, Fumio;Shimizu, Hiroshi
背景酿酒酵母（Saccharomyces cerevisiae）是工业生产乙醇、2，3-丁二醇（23 BD）等代谢产物的理想宿主菌。为了提高这些化合物的生产能力，抑制S.酿酒酵母将代谢流重定向到目标化学生产。本研究通过敲除线粒体丙酮酸转运蛋白基因（MPC 1）或线粒体自噬必需基因（ATG 32）来抑制S.结果两株菌的生长速度均比对照菌高1.6倍。C-13-代谢通量分析表明，这两种菌株呈现相似的通量分布，并成功地降低了50%的三羧酸循环通量相比，控制菌株。然而，细胞内代谢物池的大小是完全不同的，这表明在两种菌株中基因敲除的不同的代谢效应。在用于23 BD生产的试管培养中也观察到这种差异。与对照菌株（23.5 +/- 12.8 mg/L）相比，敲除ATG 32显示23 BD滴度（557.020.6 mg/L）增加23.6倍，而敲除MPC 1显示仅增加14.3倍（336.4 +/- 113.5 mg/L）。进一步的厌氧高密度发酵实验表明，MPC 1基因敲除后的酵母细胞比23 BD基因敲除后的酵母细胞更有利于乙醇的产生。结论线粒体转运蛋白和膜动力学的工程改造可以有效地控制线粒体代谢，提高酵母细胞质中化学物质的生产能力。
BackgroundSaccharomyces cerevisiae is a suitable host for the industrial production of pyruvate-derived chemicals such as ethanol and 2,3-butanediol (23BD). For the improvement of the productivity of these chemicals, it is essential to suppress the unnecessary pyruvate consumption in S. cerevisiae to redirect the metabolic flux toward the target chemical production. In this study, mitochondrial pyruvate transporter gene (MPC1) or the essential gene for mitophagy (ATG32) was knocked-out to repress the mitochondrial metabolism and improve the production of pyruvate-derived chemical in S. cerevisiae.ResultsThe growth rates of both aforementioned strains were 1.6-fold higher than that of the control strain. C-13-metabolic flux analysis revealed that both strains presented similar flux distributions and successfully decreased the tricarboxylic acid cycle fluxes by 50% compared to the control strain. Nevertheless, the intracellular metabolite pool sizes were completely different, suggesting distinct metabolic effects of gene knockouts in both strains. This difference was also observed in the test-tube culture for 23BD production. Knockout of ATG32 revealed a 23.6-fold increase in 23BD titer (557.020.6 mg/L) compared to the control strain (23.5 +/- 12.8 mg/L), whereas the knockout of MPC1 revealed only 14.3-fold increase (336.4 +/- 113.5 mg/L). Further investigation using the anaerobic high-density fermentation test revealed that the MPC1 knockout was more effective for ethanol production than the 23BD production.Conclusion These results suggest that the engineering of the mitochondrial transporters and membrane dynamics were effective in controlling the mitochondrial metabolism to improve the productivities of chemicals in yeast cytosol.