Secondary Alcohol Kinetic Resolution By G.oxydans And Characterization Of Relative Enzymes

Gluconobacter oxydans strains could incomplete oxidize substrates like sugars, alcohols to aldehydes, ketones and acids correspondingly which make it widely used in industrial applications. Based on the former research, catalyzing substrate mainly consisted of polyol sugars and primary alcohols and membrane-bound dehydrogenase had been in hot spot study. From genome sequence analysis, many kinds of oxidoreductase in G.oxydans have never been determined in resolutions of secondary alcohols.In this work, we firstly certificate the rest cells of G.oxydans could catalyze the kinetic resolutions of secondary alcohols which the secondary alcohols with single configuration were obtained. pH values in reactions were regulated which resulted in secondary alcohols with different configuration.1-phenylethanol,1-phenyl-2-propanol and 4-phenyl-2-butanol were chosen as the substrate. By analyzing the resolution results, rest cells preferred the secondary alcohols with R-configuration to S-configuration when pH was controlled at 5.0. However, when pH was at 8.0, the same rest cells preferred oppositely. After regulating the reactions, the e.e values of 1-phenylethanol,1-phenyl-2-propanol and 4-phenyl-2-butanol could reach 90%(S),100%(S) and 100%(R).Based on our research, gox2036 and gox0525 which belongs to short-chain alcohol dehydrogenase with unknown function were cloned and expressed in E.coli. Experiments about resolution of 1-phenylethanol,1-phenyl-2-propanol and 4-phenyl-2-butanol were performed and showed Gox2036 could oxidize secondary alcohols with S-configuration which R-configuration was retained. The enzymatic characterization of Gox0525 and Gox2036 were also studied in this paper.