4HFJ

X-ray Crystal Structure of a Double Bond Reductase from Nicotiana tabacum

Summary for 4HFJ

• Entry DOI: 10.2210/pdb4hfj/pdb
• Related: 4HFM 4HFN
• Descriptor: Allyl alcohol dehydrogenase (2 entities in total)
• Functional Keywords: rossmann fold, twisted b-barrel, alkene reduction, oxidoreductase
• Biological source: Nicotiana tabacum (American tobacco,tobacco)
• Total number of polymer chains: 2
• Total formula weight: 78404.30

Authors

Toogood, H.S.,Scrutton, N.S. (deposition date: 2012-10-05, release date: 2013-01-30, Last modification date: 2023-09-20)

Primary citation

Mansell, D.J.,Toogood, H.S.,Waller, J.,Hughes, J.M.,Levy, C.W.,Gardiner, J.M.,Scrutton, N.S.
Biocatalytic Asymmetric Alkene Reduction: Crystal Structure and Characterization of a Double Bond Reductase fromNicotiana tabacum.
ACS Catal, 3:370-379, 2013

PubMed Abstract: 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 (NtDBR), which belongs to the leukotriene B 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 α,β-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 ()-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 ()-bound] NtDBR complexes. These will underpin structure-driven site-saturated mutagenesis studies aimed at enhancing the reactivity, stereochemistry, and specificity of this enzyme.

PubMed: 27547488
DOI: 10.1021/cs300709m

Experimental method

X-RAY DIFFRACTION (2 Å)