期刊
CHEMBIOCHEM
卷 20, 期 2, 页码 276-281出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.201800556
关键词
alcohols; aldehydes; enzyme engineering; oxidases; thermostability
资金
- European Union [613849]
Structure-guided directed evolution of choline oxidase has been carried out by using the oxidation of hexan-1-ol to hexanal as the target reaction. A six-amino-acid variant was identified with a 20-fold increased k(cat) compared to that of the wild-type enzyme. This variant enabled the oxidation of 10 mm hexanol to hexanal in less than 24 h with 100 % conversion. Furthermore, this variant showed a marked increase in thermostability with a corresponding increase in T-m of 20 degrees C. Improved solvent tolerance was demonstrated with organic solvents including ethyl acetate, heptane and cyclohexane, thereby enabling improved conversions to the aldehyde by up to 30 % above conversion for the solvent-free system. Despite the evolution of choline oxidase towards hexan-1-ol, this new variant also showed increased specific activities (by up to 100-fold) for around 50 primary aliphatic, unsaturated, branched, cyclic, benzylic and halogenated alcohols.
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