1. An Engineered Alcohol Oxidase for the Oxidation of Primary Alcohols
- Author
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Nicholas J. Turner, William R. Birmingham, Laurent Daviet, Rachel S. Heath, Matthew P. Thompson, and Andreas Taglieber
- Subjects
Models, Molecular ,Ethyl acetate ,Protein Engineering ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Aldehyde ,Hexanal ,chemistry.chemical_compound ,Colletotrichum ,Organic chemistry ,Molecular Biology ,Thermostability ,chemistry.chemical_classification ,Molecular Structure ,010405 organic chemistry ,Organic Chemistry ,Choline oxidase ,0104 chemical sciences ,Alcohol oxidase ,Solvent ,Alcohol Oxidoreductases ,chemistry ,Alcohols ,Molecular Medicine ,Oxidation-Reduction ,Hexanol - Abstract
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 kcat 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 Tm of 20 °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.
- Published
- 2018