Structures of Alcohol Dehydrogenases from Ralstonia and Sphingobium spp. Reveal the Molecular Basis for Their Recognition of ‘Bulky–Bulky’ Ketones
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Alcohol dehydrogenases (ADHs) are applied in industrial synthetic chemistry for the production of optically active secondary alcohols. However, the substrate spectrum of many ADHs is narrow, and few, for example, are suitable for the reduction of prochiral ketones in which the carbonyl group is bounded by two bulky and/or hydrophobic groups; so-called ‘bulky–bulky’ ketones. Recently two ADHs, RasADH from Ralstonia sp. DSM 6428, and SyADH from Sphingobium yanoikuyae DSM 6900, have been described, which are distinguished by their ability to accept bulky–bulky ketones as substrates. In order to examine the molecular basis of the recognition of these substrates the structures of the native and NADPH complex of RasADH, and the NADPH complex of SyADH have been determined and refined to resolutions of 1.5, 2.9 and 2.5 Å, respectively. The structures reveal hydrophobic active site tunnels near the surface of the enzymes that are well-suited to the recognition of large hydrophobic substrates, as determined by modelling of the bulky–bulky substrate n-pentyl phenyl ketone. The structures also reveal the bases for NADPH specificity and (S)-stereoselectivity in each of the biocatalysts for n-pentyl phenyl ketone and related substrates.
KeywordsAlcohol dehydrogenase Ketoreductase NADPH Oxidoreductase Enzyme structure
This research was supported by a Marie Curie Network for Initial Training fellowships to K.K., J.K. and A.F. in the project BIOTRAINS (FP7-PEOPLE-ITN-2008-238531). We would also like to thank Prof. Wolfgang Kroutil of the University of Graz for genes encoding both RasADH and SyADH.
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