Journal Article

PreJuSER-4648

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Uneven Twins: Comparison of two enantiocomplementary hydroxynitrile lyases with a/ß--hydrolase fold

Guterl, K. ; Andexer, J. N. ; Sehl, T. ; von Langermann, J. ; Frindi-Wosch, I. ; Rosenkranz, T.FZJ* ; Fitter, J.FZJ* ; Gruber, K. ; Kragl, U. ; Eggert, T. ; Pohl, M.FZJ*

2009
Elsevier Science Amsterdam [u.a.]

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Please use a persistent id in citations: doi:10.1016/j.jbiotec.2009.03.010

Abstract: Hydroxynitrile lyases (HNLs) are applied in technical processes for the synthesis of chiral cyanohydrins. Here we describe the thorough characterization of the recently discovered R-hydroxynitrile lyase from Arabidopsis thaliana and its S-selective counterpart from Manihot esculenta (MeHNL) concerning their properties relevant for technical applications. The results are compared to available data of the structurally related S-HNL from Hevea brasiliensis (HbHNL), which is frequently applied in technical processes. Whereas substrate ranges are highly similar for all three enzymes, the stability of MeHNL with respect to higher temperature and low pH-values is superior to the other HNLs with alpha/beta-hydrolase fold. This enhanced stability is supposed to be due to the ability of MeHNL to form tetramers in solution, while HbHNL and AtHNL are dimers. The different inactivation pathways, deduced by means of circular dichroism, tryptophan fluorescence and static light scattering further support these results. Our data suggest different possibilities to stabilize MeHNL and AtHNL for technical applications: whereas the application of crude cell extracts is appropriate for MeHNL, AtHNL is stabilized by addition of polyols. In addition, the molecular reason for the inhibition of MeHNL and HbHNL by acetate could be elucidated, whereas no such inhibition was observed with AtHNL.

Keyword(s): Acetonitriles: metabolism (MeSH) ; Aldehyde-Lyases: chemistry (MeSH) ; Aldehyde-Lyases: genetics (MeSH) ; Aldehyde-Lyases: metabolism (MeSH) ; Amino Acid Sequence (MeSH) ; Arabidopsis: enzymology (MeSH) ; Arabidopsis: genetics (MeSH) ; Enzyme Stability (MeSH) ; Escherichia coli: genetics (MeSH) ; Hevea: enzymology (MeSH) ; Hevea: genetics (MeSH) ; Hydrogen-Ion Concentration (MeSH) ; Hydrolases: genetics (MeSH) ; Manihot: enzymology (MeSH) ; Manihot: genetics (MeSH) ; Molecular Sequence Data (MeSH) ; Plant Proteins: genetics (MeSH) ; Plant Proteins: metabolism (MeSH) ; Recombinant Proteins: genetics (MeSH) ; Recombinant Proteins: metabolism (MeSH) ; Stereoisomerism (MeSH) ; Substrate Specificity (MeSH) ; Temperature (MeSH) ; Time Factors (MeSH) ; Acetonitriles ; Plant Proteins ; Recombinant Proteins ; mandelonitrile ; Hydrolases ; Aldehyde-Lyases ; J ; Asymmetric carboligation (auto) ; Cyanohydrins (auto) ; Enzyme catalysis (auto) ; Enzyme stability (auto) ; Oxynitrilase (auto)

Note: The authors thank Julich Chiral Solutions/Codexis for providing MeHNL-DNA and MeHNL-Cys81Ala expressing E. coli cells, Astrid Wirtz for HPLC measurements and Sabine Kruschinski for technical assistance. This work was partially supported by the BMBF in frame of project "Biokatalytische Hydrocyanierung & Hydroformylierung (BioHydroForm) FKZ 0313402C" and by the Deutsche Forschungsgemeinschaft in frame of the research training group "BioNoCo" GK 1166.

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Contributing Institute(s):

1. Molekulare Biophysik (ISB-2)

Research Program(s):

1. Programm Biosoft (N03)

Appears in the scientific report 2009

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