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Optimization of Alcohol Dehydrogenase for Industrial Scale Oxidation of Lactols.
Biotechnology Journal ( IF 3.2 ) Pub Date : 2020-08-26 , DOI: 10.1002/biot.202000171
Sebastian Bartsch 1 , Jan Brummund 2 , Sabrina Köpke 1 , Harrie Straatman 2 , Andreas Vogel 1 , Martin Schürmann 2
Affiliation  

Alcohol dehydrogenases (ADH) are widely used to enantioselectively reduce ketones to chiral alcohols, but their application in industrial scale oxidations is rare. Reasons are the need for an NAD(P)+ cofactor regeneration system, often low performance in oxidative reactions and the limited substrate scope of ADHs. ADHA from Candida magnoliae DSMZ 70638 is identified to efficiently catalyze the regio‐selective hydroxy‐lactone oxidations to hydroxy‐lactones. Hydroxy‐lactones are common intermediates in industrial processes to cholesterol lowering (va)statin drugs. A biocatalytic aliphatic hydroxy‐lactone oxidation process is developed using pure oxygen as oxidant reaching volumetric productivities of up to 12 g L−1 h−1, product concentrations of almost 50 g L−1 and 95% reaction yield. For co‐factor recycling a previously engineered, water‐forming NAD(P)H‐oxidase from Streptococcus mutans is used. The process is scaled up to industrial pilot plant scale and it could be demonstrated that ADH catalyzed oxidations can be developed to efficient and safe processes. However, the ADHA wild‐type enzyme is not productive enough in chlorolactol oxidation. Therefore, enzyme engineering and multi‐parameter screening is successfully applied to optimize the enzyme for the target reaction. The optimized ADHA variant shows a 17‐fold higher oxidative activity, a 26°C increased stability and is applied to develop an efficient chlorolactol oxidation process.

中文翻译:

用于工业规模氧化乳糖的醇脱氢酶的优化。

醇脱氢酶(ADH)广泛用于将对映体选择性还原为手性醇,但是在工业规模的氧化中却很少使用。原因是需要NAD(P)+辅因子再生系统,在氧化反应中通常性能低下,而ADH的底物范围有限。玉兰假丝酵母DSMZ 70638的ADHA可有效催化区域选择性羟基内酯氧化为羟基内酯。羟基内酯是工业过程中降低胆固醇(va)他汀类药物的常见中间体。使用纯氧作为氧化剂,开发了一种生物催化的脂肪族羟基内酯氧化工艺,其容积生产率高达12 g L -1 h -1,产物浓度几乎为50 g L -1,反应产率为95%。为了进行辅因子回收,使用了以前设计的,来自变形链球菌的水形成NAD(P)H-氧化酶。该方法已扩大到工业中试规模,可以证明ADH催化的氧化技术可以发展为高效安全的方法。但是,ADHA野生型酶在氯丙醇氧化中的生产力不足。因此,酶工程和多参数筛选已成功应用于优化目标反应的酶。经过优化的ADHA变体显示出高17倍的氧化活性,稳定性提高了26°C,可用于开发有效的氯乳酸氧化工艺。
更新日期:2020-08-26
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