Productive Amyrin Synthases for Efficient α-Amyrin Synthesis in Engineered Saccharomyces cerevisiae,ACS Synthetic Biology

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Productive Amyrin Synthases for Efficient α-Amyrin Synthesis in Engineered Saccharomyces cerevisiae
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2018-09-14 00:00:00 , DOI: 10.1021/acssynbio.8b00176
Yuan Yu _{1,

2,

3} , Pengcheng Chang ₂ , Huan Yu ₄ , Huiyong Ren ₄ , Danning Hong ₄ , Zeyan Li ₄ , Ying Wang ₂ , Hao Song ₁ , Yixin Huo ₄ , Chun Li _{1,

2}

Affiliation

α-Amyrin is a plant-derived pentacyclic triterpenoid, with a lot of important physiological and pharmacological activities. The formation of α-amyrin from (3S)-2,3-oxidosqualene is catalyzed by α-amyrin synthase (α-AS), a member of the oxidosqualene cyclase (OSC) protein family. However, α-amyrin is not yet commercially developed due to its extremely low productivity in plants. The engineered Saccharomyces cerevisiae with efficient α-amyrin production pathway could be used as an alternative and sustainable solution to produce α-amyrin from renewable raw materials. To efficiently improve α-amyrin production in S. cerevisiae, we identified two α-ASs, EjAS and MdOSC1 from Eriobotrya japonica and Malus × domestica, respectively, through strict bioinformatics screening criteria and phylogenetic analysis. The specific activities of purified EjAS and MdOSC1 were 0.0032 and 0.0293 μmol/min/mg, respectively. EjAS produced α-amyrin and β-amyrin at a ratio of 17:3, MdOSC1 produced α-amyrin, β-amyrin and lupeol at a ratio of 86:13:1, indicating MdOSC1 had significantly higher specific activity and higher ratio of α-amyrin than EjAS. Furthermore, MdOSC1 was introduced into S. cerevisiae combining with the increased supply of (3S)-2,3-oxidosqualene to achieve the encouraging α-amyrin production, and the titer of α-amyrin achieved 11.97 ± 0.61 mg/L, 5.8 folds of the maximum production reported.

中文翻译：

高效工程合成酿酒酵母中高效生产α-淀粉状蛋白的生产性淀粉样蛋白合酶

α-Amyrin是植物来源的五环三萜，具有许多重要的生理和药理活性。由（3 S）-2,3-氧化角鲨烯形成α-淀粉酶是由氧化角鲨烯环化酶（OSC）蛋白家族成员α-淀粉酶合成酶（α-AS）催化的。然而，由于其在植物中的极低的生产率，α-胰岛淀粉蛋白尚未被商业开发。具有高效α-amyrin生产途径的工程酿酒酵母可以用作从可再生原料生产α-amyrin的替代且可持续的解决方案。为了有效提高酿酒酵母中α-amyrin的产量，我们鉴定了两种来自日本Eriobotrya japonica和Malus×domestica的α-AS，EjAS和MdOSC1。分别通过严格的生物信息学筛选标准和系统发育分析。纯化的EjAS和MdOSC1的比活分别为0.0032和0.0293μmol/ min / mg。EjAS以17：3的比例生成α-amyrin和β-amyrin，MdOSC1以86：13：1的比例生成α-amyrin，β-amyrin和lupeol，表明MdOSC1具有明显更高的比活和更高的α比-amyrin比EjAS。此外，将MdOSC1引入酿酒酵母中，同时增加了（3 S）-2,3-氧化角鲨烯的供应，从而实现了令人鼓舞的α-amyrin生产，α-amyrin的效价达到11.97±0.61 mg / L，5.8报道的最大产量的几倍。

更新日期：2018-09-14

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