Pushing the limits: Cyclodextrin-based intensification of bioreductions

Area of Science:

• Biocatalysis and Enzyme Engineering
• Organic Synthesis
• Green Chemistry

Background:

• Asymmetric reduction of ketones to chiral alcohols is crucial in synthesis.
• Biocatalysts offer high enantiopurity but often suffer from low reaction output due to limited enzyme stability.
• Previous strategies used hexane as a second phase to protect catalysts.

Purpose of the Study:

• To enhance the efficiency and stability of biocatalytic reduction of ketones.
• To investigate the effect of 2-hydroxypropyl-β-cyclodextrin on biocatalyst performance.
• To achieve high product concentrations and enantiopurity in bioreductions.

Main Methods:

• Employing an E. coli strain co-expressing xylose reductase and formate dehydrogenase for NADH recycling.
• Utilizing 2-hydroxypropyl-β-cyclodextrin as an additive in the bioreduction of o-chloroacetophenone.
• Comparing the performance with and without cyclodextrin, and with hexane as a control.
• Investigating the role of a D51A mutant of xylose reductase.

Main Results:

• Addition of 2-hydroxypropyl-β-cyclodextrin enabled product concentrations up to 29% w/v with 99.97% enantiomeric excess (e.e.).
• The cyclodextrin stabilized and activated the biocatalyst, with activity observed both free and associated with biomass.
• Substitution of wild-type xylose reductase with a D51A mutant further improved bioreduction.
• Replacing hexane with 2-hydroxypropyl-β-cyclodextrin increased the yield on biocatalyst 6.3-fold.

Conclusions:

• 2-hydroxypropyl-β-cyclodextrin significantly enhances biocatalytic ketone reduction, improving product concentration, enantiopurity, and catalyst stability.
• This cyclodextrin-mediated enhancement is potentially applicable across various enzyme classes, catalyst forms, and substrates.
• Cyclodextrins represent a promising alternative to traditional phase-transfer methods for improving biocatalytic processes.