Mevalonate is a key precursor in isoprenoid biosynthesis and a promising commodity chemical. Although mevalonate is a native metabolite in Saccharomyces cerevisiae, its production is challenged by the relatively low flux toward acetyl-CoA in this yeast. In this study we explore different approaches to increase acetyl-CoA supply in S. cerevisiae to boost mevalonate production. Stable integration of a feedback-insensitive acetyl-CoA synthetase (Se-acsL641P) from Salmonella enterica and the mevalonate pathway from Enterococcus faecalis results in the production of 1,390 ± 10 mg/l of mevalonate from glucose. While bifid shunt enzymes failed to improve titers in high-producing strains, inhibition of squalene synthase (ERG9) results in a significant enhancement. Finally, increasing coenzyme A (CoA) biosynthesis by overexpression of pantothenate kinase (CAB1) and pantothenate supplementation further increased production to 3,830 ± 120 mg/l. Using strains that combine these strategies in lab-scale bioreactors results in the production of 13.3 ± 0.5 g/l, which is ∼360-fold higher than previously reported mevalonate titers in yeast. This study demonstrates the feasibility of engineering S. cerevisiae for high-level mevalonate production.

中文翻译：

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工程乙酰辅酶A供应和ERG9抑制以增强酿酒酵母中甲羟戊酸的产生

甲羟戊酸是类异戊二烯生物合成的关键前体，是一种很有前景的商品化学品。尽管甲羟戊酸是酿酒酵母中的天然代谢物，但它的生产受到这种酵母中乙酰辅酶A 相对较低的通量的挑战。在这项研究中，我们探索了增加酿酒酵母中乙酰辅酶A供应以促进甲羟戊酸生产的不同方法。来自肠沙门氏菌的反馈不敏感的乙酰辅酶 A 合成酶 (Se-acsL641P) 和来自粪肠球菌的甲羟戊酸途径的稳定整合导致从葡萄糖中产生 1,390 ± 10 mg/l 的甲羟戊酸。虽然双歧分流酶未能提高高产菌株的滴度，但抑制角鲨烯合酶 (ERG9) 会导致显着增强。最后，通过泛酸激酶 (CAB1) 的过度表达和泛酸补充剂增加辅酶 A (CoA) 的生物合成，进一步将产量提高到 3,830 ± 120 mg/l。在实验室规模的生物反应器中使用结合这些策略的菌株可产生 13.3 ± 0.5 g/l，这比之前报道的酵母中甲羟戊酸滴度高 360 倍。这项研究证明了工程化酿酒酵母用于高水平甲羟戊酸生产的可行性。