Asymmetric reduction of ketones and β-keto esters by (S)-1-phenylethanol dehydrogenase from denitrifying bacterium Aromatoleum aromaticum

Abstract

Enzyme-catalyzed enantioselective reductions of ketones and keto esters have become popular for the production of homochiral building blocks which are valuable synthons for the preparation of biologically active compounds at industrial scale. Among many kinds of biocatalysts, dehydrogenases/reductases from various microorganisms have been used to prepare optically pure enantiomers from carbonyl compounds. (S)-1-phenylethanol dehydrogenase (PEDH) was found in the denitrifying bacterium Aromatoleum aromaticum (strain EbN1) and belongs to the short-chain dehydrogenase/reductase family. It catalyzes the stereospecific oxidation of (S)-1-phenylethanol to acetophenone during anaerobic ethylbenzene mineralization, but also the reverse reaction, i.e., NADH-dependent enantioselective reduction of acetophenone to (S)-1-phenylethanol. In this work, we present the application of PEDH for asymmetric reduction of 42 prochiral ketones and 11 β-keto esters to enantiopure secondary alcohols. The high enantioselectivity of the reaction is explained by docking experiments and analysis of the interaction and binding energies of the theoretical enzyme-substrate complexes leading to the respective (S)- or (R)-alcohols. The conversions were carried out in a batch reactor using Escherichia coli cells with heterologously produced PEDH as whole-cell catalysts and isopropanol as reaction solvent and cosubstrate for NADH recovery. Ketones were converted to the respective secondary alcohols with excellent enantiomeric excesses and high productivities. Moreover, the progress of product formation was studied for nine para-substituted acetophenone derivatives and described by neural network models, which allow to predict reactor behavior and provides insight on enzyme reactivity. Finally, equilibrium constants for conversion of these substrates were derived from the progress curves of the reactions. The obtained values matched very well with theoretical predictions.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-014-6309-z) contains supplementary material, which is available to authorized users.

Molecular Genetic and Crystal Structural Analysis of 1-(4-Hydroxyphenyl)-Ethanol Dehydrogenase from ‘Aromatoleum aromaticum' EbN1

“…Furthermore, results from the crystal structure analyses corroborated the potential of these dehydrogenases for producing enantiopure secondary alcohols from a wide range of prochiral ketones and esters as recently demonstrated for Ped from 'A. aromaticum' EbN1 [Breuer et al, 2008;Dudzik et al, 2015;Gröger et al, 2006;Hummel et al, 1999]. Conversely, only one enantiomer can be selectively oxidized from a racemic mixture of an alcohol.…”

DFT-based prediction of reactivity of short-chain alcohol dehydrogenase

“…Such protonation was also confirmed in previous docking studies both for PEDH and Hped [8, 39]. Moreover, our QM calculations (data not shown) demonstrated, that if TyrO − /Lys-NH 3 + model is formed with ketone, TyrO − spontaneously strips proton from O2′ ribose and proton from Lys-NH 3 + is transferred on O2′ atom, resulting with formation of TyrOH/Lys-NH 2 system.…”

ANN Architecture Specifications for Modelling of Open-Cell Aluminum under Compression

“…Nevertheless, including the problematic results leaves potential designers on the safe side. e solution to this issue was based on the approach proposed in [42] and will be described in…”

“…Again, considerable initial RE i s might be attributed to the fact that, in case of the beginning of the data scope, denominators in relative errors are very small and that prototype samples might have behaved irregularly due to structural imperfections and this region is mechanically more sensitive to such an impact. As has been already mentioned, the solution for the described issue of the initial region was inspired by the study [42] and separate networks were trained for the initial stress-strain curve region. Again, this was performed only for sample 8, as an example.…”