Asymmetric Reduction of Activated Alkenes by Pentaerythritol Tetranitrate Reductase: Specificity and Control of Stereochemical Outcome by Reaction Optimisation

Abstract

We show that pentaerythritol tetranitrate reductase (PETNR), a member of the 'ene' reductase old yellow enzyme family, catalyses the asymmetric reduction of a variety of industrially relevant activated alpha,beta-unsaturated alkenes including enones, enals, maleimides and nitroalkenes. We have rationalised the broad substrate specificity and stereochemical outcome of these reductions by reference to molecular models of enzyme-substrate complexes based on the crystal complex of the PETNR with 2-cyclohexenone 4a. The optical purity of products is variable (49-99% ee), depending on the substrate type and nature of substituents. Generally, high enantioselectivity was observed for reaction products with stereogenic centres at Cbeta (>99% ee). However, for the substrates existing in two isomeric forms (e.g., citral 11a or nitroalkenes 18-19a), an enantiodivergent course of the reduction of E/Z-forms may lead to lower enantiopurities of the products. We also demonstrate that the poor optical purity obtained for products with stereogenic centres at Calpha is due to non-enzymatic racemisation. In reactions with ketoisophorone 3a we show that product racemisation is prevented through reaction optimisation, specifically by shortening reaction time and through control of solution pH. We suggest this as a general strategy for improved recovery of optically pure products with other biocatalytic conversions where there is potential for product racemisation.

An enoate reductase Achr-OYE4 from Achromobacter sp. JA81: characterization and application in asymmetric bioreduction of C=C bonds

“…The α-substituted nitroalkene 4a was also accepted as a substrate by Achr-OYE4, but the resulting product was racemic (Table 3), which was mainly the result of spontaneous racemization of the stereogenic center at Cα in aqueous media, as reported earlier (Fryszkowska et al 2009;Hall et al 2007). Substrate 4a with a methyl substituent at the α-position preferentially interacted with the enzyme over 3a, with the methyl substituent at the β-position, as judged by the higher conversion of 4a.…”

Identification and characterization of a novel “thermophilic-like” Old Yellow Enzyme from the genome of Chryseobacterium sp. CA49

“…As organic solvents are often used to increase substrate solubility or product enantiopurity [34,35] and even improve the catalytic property of enzyme [36], we investigated the solvent stability of Chr-OYE3 in the presence of several solvents (both water-miscible and -immiscible) of varied log P (from −1.49 to 2.52) with a volumetric ratio of 5-80% (Fig. 8).…”

Old Yellow Enzyme-Catalysed Asymmetric Hydrogenation: Linking Family Roots with Improved Catalysis

“…Catalysts 2017, 7, 130 2 of 24 activated alkenes with an electron withdrawing group (EWG) such as aldehyde, ketone, anhydride [8,10,11], nitro [9,12,13], (di)ester [8,[14][15][16][17], (di)carboxylic acid [18][19][20], cyclic imide [21][22][23], nitrile [24], β-cyanoacrylate [25], β-nitroacrylate [26], and several other functional groups [27]. There are many examples of the high industrial potential of OYEs for the synthesis of valuable target products [28][29][30][31].…”