Synthesis of (R)-mandelic acid and (R)-mandelic acid amide by recombinant E. coli strains expressing a (R)-specific oxynitrilase and an arylacetonitrilase

Objectives: Chiral 2-hydroxycarboxylic acids and 2-hydroxycarboxamides are valuable synthons for the chemical industry.

Results: The biocatalytic syntheses of (R)-mandelic acid and (R)-mandelic acid amide by recombinant Escherichia coli clones were studied. Strains were constructed which simultaneously expressed a (R)-specific oxynitrilase (hydroxynitrile lyase) from the plant Arabidopsis thaliana together with the arylacetonitrilase from the bacterium Pseudomonas fluorescens EBC191. In addition, recombinant strains were constructed which expressed a previously described acid tolerant variant of the oxynitrilase and an amide forming variant of the nitrilase. The whole cell catalysts which simultaneously expressed the (R)-specific oxynitrilase and the wild-type nitrilase transformed in slightly acidic buffer systems benzaldehyde plus cyanide preferentially to (R)-mandelic acid with ee-values > 95%. The combination of the (R)-specific oxynitrilase with the amide forming nitrilase variant gave whole cell catalysts which converted at pH-values ≤ pH 5 benzaldehyde plus cyanide with a high degree of enantioselectivity (ee > 90%) to (R)-mandelic acid amide. The acid and the amide forming catalysts also converted chlorinated benzaldehydes with cyanide to chlorinated mandelic acid or chlorinated mandelic acid amides.

Conclusions: Efficient systems for the biocatalytic production of (R)-2-hydroxycarboxylic acids and (R)-2-hydroxycarboxamides were generated.

Keywords: Biotransformations; Chiral synthesis; Enzyme cascades; Hydroxynitrile lyase; Nitrilase; Oxynitrilase.