Synthesis of ethyl ( R )-mandelate using recombinant Carboxydothermus hydrogenoformans alcohol dehydrogenase produced by two yeast species

Yeast cell catalysts carrying a recombinant Carboxydothermus hydrogenoformans alcohol dehydrogenase (ChADH) gene were used to synthesise ethyl ( R )-mandelate. Transgenic Arxula adeninivorans and Hansenula polymorpha strains were constructed to produce recombinant ChADH at high concentrations. Biochemical parameters such as pH and temperature optima, thermostability and substrate specificity were determined for the enzyme synthesized in Arxula adeninivorans . The recombinant enzyme combined with a substrate-coupled cofactor regeneration system and permeabilized Arxula adeninivorans and Hansenula polymorpha cell catalysts co-expressing ChADH and Bacillus megaterium glucose dehydrogenase ( BmGDH ) for enzyme-coupled cofactor regeneration, were used to synthesize ethyl (R)-mandelate. Comparison of purified recombinant ChADH/BmGDH and both of the yeast cell based catalysts ability to synthesize ethyl ( R )-mandelate demonstrated that Hansenula polymorpha cell catalysts were able to produce the highest yield ( Hansenula polymorpha 6.07 mmol l −1  h −1 ) while Arxula adeninivorans produced approximately half this amount (3.07 mmol l −1  h −1 ). The maximum conversion achieved was 98% with a high enantiomeric excess (>98%).