Journal Article

PreJuSER-23196

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Hydroxynitrile lyases with alpha/beta-hydrolase fold: two enzymes with almost identical 3D structures but opposite enantioselectivities and different reaction mechanisms

Andexer, J. N. ; Staunig, N. ; Eggert, T. ; Kratky, C. ; Pohl, M.FZJ* ; Gruber, K.

2012
Wiley-VCH Weinheim

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Please use a persistent id in citations: doi:10.1002/cbic.201200239

Abstract: Hydroxynitrile lyases (HNLs) catalyze the cleavage of cyanohydrins to yield hydrocyanic acid (HCN) and the respective carbonyl compound and are key enzymes in the process of cyanogenesis in plants. In organic syntheses, HNLs are used as biocatalysts for the formation of enantiopure cyanohydrins. We determined the structure of the recently identified, R-selective HNL from Arabidopsis thaliana (AtHNL) at a crystallographic resolution of 2.5 Å. The structure exhibits an α/β-hydrolase fold, very similar to the homologous, but S-selective, HNL from Hevea brasiliensis (HbHNL). The similarities also extend to the active sites of these enzymes, with a Ser-His-Asp catalytic triad present in all three cases. In order to elucidate the mode of substrate binding and to understand the unexpected opposite enantioselectivity of AtHNL, complexes of the enzyme with both (R)- and (S)-mandelonitrile were modeled using molecular docking simulations. Compared to the complex of HbHNL with (S)-mandelonitrile, the calculations produced an approximate mirror image binding mode of the substrate with the phenyl rings located at very similar positions, but with the cyano groups pointing in opposite directions. A catalytic mechanism for AtHNL is proposed, in which His236 from the catalytic triad acts as a general base and the emerging negative charge on the cyano group is stabilized by main-chain amide groups and an α-helix dipole very similar to α/β-hydrolases. This mechanistic proposal is additionally supported by mutagenesis studies.

Keyword(s): J ; biocatalysis (auto) ; C-C lyase (auto) ; enzyme mechanisms (auto) ; molecular modeling (auto) ; X-ray crystallography (auto)

Note: We acknowledge financial support from the Austrian Science Foundation (FWF) through projects P17132 (to C.K.) and L148 (to K.G.). X-ray diffraction data were collected at the EMBL/DESY beamline X13 in Hamburg, Germany and we are indebted to the beamline staff for their help. For her help with enzyme purification, we thank Ilona Frindi-Wosch.

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Contributing Institute(s):

1. Biotechnologie 2 (IBT-2)

Research Program(s):

1. Biotechnologie (PBT)

Appears in the scientific report 2012

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Database coverage:
; BIOSIS Previews ; Current Contents - Life Sciences ; JCR ; NCBI Molecular Biology Database ; SCOPUS ; Science Citation Index ; Science Citation Index Expanded ; Thomson Reuters Master Journal List ; Web of Science Core Collection

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