초록
<P><B>Abstract</B></P> <P>Ketoreductase <I>Ch</I>KRED12 from <I>Chryseobacterium</I> sp. CA49 was identified as a highly stereoselective catalyst for the bioreductive production of (2<I>S</I>,3<I>S</I>)-ethyl 3-hydroxy-2-methoxy-3-phenylpropanoate (EMHP) through dynamic kinetic resolution. Its performance was further improved using semi-rational design strategies targeting residues Gln151 and Met191. Mutant M191S displayed enhanced activity (4-fold), producing (2<I>S</I>,3<I>S</I>)-EMHP with >99% ee and > 99/1 dr; two double mutants (Q151F/M191 L and Q151Y/M191 L) displayed reversed stereoselectivity and enhanced activity (~3-fold), producing (2<I>R</I>,3<I>R</I>)-EMHP with >99% ee and 98/2 dr. Models of the mutants docked with the substrate and NAD<SUP>+</SUP> revealed possible mechanism of the improved characteristics.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Ketoreductase <I>Ch</I>KRED12 was used to reduce 2-methoxy-3-oxo-3- phenylpropanoate. </LI> <LI> Semi-rational design was performed to improve the activity and selectivity. </LI> <LI> Mutant M191S produced (2<I>S</I>,3<I>S</I>)-EMHP with >99% ee and > 99/1 dr. </LI> <LI> Two double mutants produced (2<I>R</I>,3<I>R</I>)-EMHP with >99% ee and 98/2 dr. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>