Immobilization conditions of ketoreductase on enantioselective reduction in a gas-solid bioreactor.
The immobilization conditions of commercial ketoreductase for continuous enantioselective reduction in the gas-phase reaction were investigated with respect to the immobilization efficiency (residual activity and protein loading) and the gas-phase reaction efficiency (initial reaction rate, half-life, and enantioselectivity). For the analyses, ketoreductase was first immobilized by physical deposition on glass supports and the reduction of 2-butanone to (S)-2-butanol with the concomitant regeneration of NADH by 2-propanol was used as a model reaction. The optimal conditions of enzyme immobilization were obtained using an absolute pressure of 100 hPa for drying, a pH between 6.5 and 7.0, and a buffer concentration of 50 mM. The buffer concentration in particular had a strong effect on both the enzyme activity and enantioselectivity. Under optimal immobilization conditions, the thermostability of ketoreductase in the gas-phase system was enhanced compared to the aqueous-phase system, while the enantioselectivity was successfully maintained at a level identical to that of the native enzyme. These results indicate that the gas-phase reaction has a great potential for industrial production of chiral compounds, but requires careful optimization of immobilization conditions for the reaction to progress effectively.
Nagayama
Kazuhito
K
Department of Materials Science and Engineering, Kochi National College of Technology, Kochi, Japan. knaga@ms.kochi-ct.ac.jp
Spiess
Antje C
AC
Büchs
Jochen
J
D019840:2-Propanol / Q000378:metabolism; D000426:Alcohol Dehydrogenase / Q000737:chemistry* / Q000378:metabolism; D019149:Bioreactors; D002021:Buffers; D002074:Butanones / Q000378:metabolism; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009243:NAD / Q000378:metabolism; D011312:Pressure; D013237:Stereoisomerism
D002021:Buffers; D002074:Butanones; D004800:Enzymes, Immobilized; D009243:NAD; C005222:methylethyl ketone; D000426:Alcohol Dehydrogenase; D019840:2-Propanol