Structure and mechanism of the ThDP-dependent benzaldehyde lyase from Pseudomonas fluorescens

Authors


  • Note
    After submission of this manuscript, we received a preprint of the following paper reporting that the mutation of His29 to alanine reduces the BAL activity to 5%. Kneen MM, Pogozheva ID, Kenyon GL & McLeish MJ (2005) Exploring the active site of benzaldehyde lyase by modeling and mutagenesis. Biochim Biophys Acta: Proteins and Proteomics, doi:10.1016/j.bbapap2005.08.025

G. E. Schulz, Institut für Organische Chemie und Biochemie, Albertstr. 21, Freiburg im Breisgau, Germany 79104
Tel: +49 761 203 6058
Fax: +49 761 203 6161
Email: georg.schulz@ocbc.uni-freiburg.de

Abstract

Pseudomonas fluorescens is able to grow on R-benzoin as the sole carbon and energy source because it harbours the enzyme benzaldehyde lyase that cleaves the acyloin linkage using thiamine diphosphate (ThDP) as a cofactor. In the reverse reaction, this lyase catalyses the carboligation of two aldehydes with high substrate and stereospecificity. The enzyme structure was determined by X-ray diffraction at 2.6 Å resolution. A structure-based comparison with other proteins showed that benzaldehyde lyase belongs to a group of closely related ThDP-dependent enzymes. The ThDP cofactors of these enzymes are fixed at their two ends in separate domains, suspending a comparatively mobile thiazolium ring between them. While the residues binding the two ends of ThDP are well conserved, the lining of the active centre pocket around the thiazolium moiety varies greatly within the group. Accounting for the known reaction chemistry, the natural substrate R-benzoin was modelled unambiguously into the active centre of the reported benzaldehyde lyase. Due to its substrate spectrum and stereospecificity, the enzyme extends the synthetic potential for carboligations appreciably.

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