期刊
ACS CATALYSIS
卷 3, 期 3, 页码 370-379出版社
AMER CHEMICAL SOC
DOI: 10.1021/cs300709m
关键词
double bond reductase; asymmetric alkene reduction; biocatalysis; crystal structure; Nicotiana tabacum
资金
- UK Biotechnology and Biological Sciences Research Council (BBSRC) [BB/E010717/1]
- Royal Society
- Engineering and Physical Sciences Research Council (EPSRC) [EP/J020192/1]
- BBSRC [BB/J015512/1, BB/E010717/1] Funding Source: UKRI
- EPSRC [EP/J020192/1] Funding Source: UKRI
The application of biocatalysis for the asymmetric reduction of activated C=C is a powerful tool for the manufacture of high-value chemical commodities. The biocatalytic potential of -ene reductases from the Old Yellow Enzyme (OYE) family of oxidoreductases is well-known; however, the specificity of these enzymes toward mainly small molecule substrates has highlighted the need to discover -ene reductases from different enzymatic classes to broaden industrial applicability. Here, we describe the characterization of a flavin-free double bond reductase from Nicotiana tabacum (NtDBR), which belongs to the leukotriene B-4. dehydrogenase (LTD) subfamily of the zinc-independent, medium chain dehydrogenase/reductase superfamily of enzymes. Using steady-state kinetics and biotransformation reactions, we have demonstrated the regio- and stereospecificity of NtDBR against a variety of alpha,beta-unsaturated activated alkenes. In addition to catalyzing the reduction of typical LTD substrates and several classical OYE-like substrates, NtDBR also exhibited complementary activity by reducing non-OYE substrates (i.e., reducing the exocyclic C=C double bond of (R)-pulegone) and in some cases showing an opposite stereopreference in comparison with the OYE family member pentaerythritol tetranitrate (PETN) reductase. This serves to augment classical OYE -ene reductase activity and, coupled with its aerobic stability, emphasizes the potential industrial value of NtDBR. Furthermore, we also report the X-ray crystal structures of the holo-, binary NADP(H)-bound, and ternary [NADP(+) and 4-hydroxy-3-methoxycinnamaldehyde (9a)-bound] NtDBR complexes. These will underpin structure-driven site-saturated mutagenesis studies aimed at enhancin g the reactivity, stereochemistry, and specificity of this enzyme.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据