Kinetic modeling of acetophenone reduction catalyzed by alcohol dehydrogenase from Thermoanaerobacter sp.

NADPH-dependent alcohol dehydrogenase (ADH) from Thermoanaerobacter sp. was kinetically characterized using reduction of acetophenone as a model. To achieve 98% conversion of acetophenone, cofactor regeneration by oxidation of 2-propanol with the same enzyme was used. The enzyme was stable in the batch reactor. It was enantioselective towards (S)-1-phenylethanol (ee>99.5%). Due to its high deactivation in continuously operated stirred tank reactor (kd=0.0141 min-1) there was no way to keep high conversion of acetophenone at 98%. The deactivation occurred in the repetitive batch as well. A mathematical model for the acetophenone reduction with cofactor regeneration describing the behaviour in a batch, repetitive-batch and continuously stirred tank reactor was developed

Continuous asymmetric ketone reduction processes with recombinant Escherichia coli

The reduction of methyl acetoacetate was carried out in continuously operated biotransformation processes catalyzed by recombinant Escherichia coli cells expressing an alcohol dehydrogenase from Lactobacillus brevis. Three different cell types were applied as biocatalysts in three different cofactor regeneration approaches. Both processes with enzyme-coupled cofactor regeneration catalyzed by formate dehydrogenase or glucose dehydrogenase are characterized by a rapid deactivation of the biocatalyst. By contrast the processes with substrate-coupled cofactor regeneration by alcohol dehydrogenase catalyzed oxidation of 2-propanol could be run over a period of 7 weeks with exceedingly high substrate and cosubstrate concentrations of up to 2.5 and 2.8 mol L(-1), respectively. Even under these extreme conditions, the applied biocatalyst showed a good stability with only marginal leakage of intracellular cofactors