Improving acetyl-CoA biosynthesis in Saccharomyces cerevisiae via the overexpression of pantothenate kinase and PDH bypass.
Improving acetyl-CoA biosynthesis in Saccharomyces cerevisiae via the overexpression of pantothenate kinase and PDH bypass.
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DOI：
10.1186/s13068-017-0726-z
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发表时间：
2017
影响因子：
6.3
通讯作者：
Jiang R
中科院分区：
文献类型：
--
作者：
Liu W;Zhang B;Jiang R
乙酰辅酶A是酿酒酵母中重要的前体物质。以前已经采用各种方法来提高其胞质水平，重点是改造乙酰辅酶A的“乙酰基”部分。据我们所知，迄今为止还没有关于工程化“-CoA”部分的报告。在这项研究中，我们试图工程化S。酿酒酵母通过泛酸激酶过表达从“-CoA”部分（来自S.酿酒酵母，CoA合成的限速酶）和通过PDH旁路引入的“乙酰基-“部分（来自S.酿酒酵母和来自肠沙门氏菌的SeAcs L 641 P）。由于乙酰辅酶A是柚皮素的前体，因此构建了一种产生柚皮素的报告菌株以反映胞质乙酰辅酶A水平。发现单独的PanK过表达或PDH旁路引入仅导致柚皮素滴度的2倍或6.74倍增加，但在0.5mM底物对香豆酸存在下，与对照（菌株CENF 09）相比，两者的组合（菌株CENF PAA 01）导致24.4倍增加。PanK底物泛酸的补充导致柚皮素产量的另外19%的增加。为了大大提高酵母胞质溶胶中的乙酰辅酶A水平，同时工程化“乙酰-”部分和“-CoA”部分是可行的。CoA供应不足可能加剧乙酰CoA短缺，导致目标产物收率降低。本文的在线版本（doi：10.1186/s13068-017-0726-z）包含补充材料，可供授权用户使用。
Acetyl-CoA is an important precursor in Saccharomyces cerevisiae. Various approaches have been adopted to improve its cytosolic level previously with the emphasis on engineering the “acetyl-” part of acetyl-CoA. To the best of our knowledge, there have been no reports on engineering the “-CoA” part so far. In this study, we had tried to engineer S. cerevisiae from both the “-CoA” part via pantothenate kinase overexpression (PanK from S. cerevisiae, the rate-limiting enzyme for CoA synthesis) and the “acetyl-“part through PDH bypass introduction (ALD6 from S. cerevisiae and SeAcs L641P from Salmonella enteric). A naringenin-producing reporter strain had been constructed to reflect cytosolic acetyl-CoA level as acetyl-CoA is the precursor of naringenin. It was found that PanK overexpression or PDH bypass introduction alone only led to a twofold or 6.74-fold increase in naringenin titer, but the combination of both (strain CENFPAA01) had resulted in 24.4-fold increase as compared to the control (strain CENF09) in the presence of 0.5 mM substrate p-coumaric acid. The supplement of PanK substrate pantothenate resulted in another 19% increase in naringenin production. To greatly enhance acetyl-CoA level in yeast cytosol, it is feasible to engineer both the “acetyl-” part and the “-CoA” part simultaneously. Insufficient CoA supply might aggravate acetyl-CoA shortage and cause low yield of target product. The online version of this article (doi:10.1186/s13068-017-0726-z) contains supplementary material, which is available to authorized users.