Flexible ring N-heterocyclic alcohols for drug discovery by enantiocomplementary ketoreductases

Abstract Bioreduction of prochiral N -(3-oxobutyl)- and N -(2-oxopropyl)heterocycles—comprising (partially) saturated, flexible rings—was explored, using microbial whole-cell ketoreductases, including selected wild-type yeast strains and enantiocomplementary recombinant alcohol dehydrogenases (ADHs). Initial screening reactions with yeast strains resulted in ( S )-alcohols with low to moderate conversions. Optimization of the reaction conditions (substrate and cosubstrate concentration) of the yeast-catalyzed bioreductions was performed using design of experiments (DOE), resulting in significant increases of conversion for most substrate–yeast strain combinations. Thus, several substrates could be fully or almost fully converted to the corresponding enantiopure ( S )-alcohols with wild-type yeasts. Extending the scope of biocatalysts to recombinant ADHs [two ( S )-selective ADHs from Candida parapsilosis (CpADH) and Rhodococcus aetherivorans (RaADH) and an ( R )-selective one from Lactobacillus kefir (LkADH)] resulted in further improvements. Of the investigated biocatalysts, CpADH proved to be the most versatile, converting the more challenging shorter sidechain N -(2-oxopropyl)heterocycles as well. The ( R )-selective Lactobacillus kefir ADH provided enantiopure ( R )-alcohols, enabling the efficient enantiocomplementary synthesis of the target alcohols. The preparative-scale bioreductions under optimal conditions usually showed superior conversions compared to the screening reactions, resulting in enantiopure ( S )- and ( R )-enantiomers (ee > 99%) of these synthetically valuable chiral fragments. Molecular docking studies with the substrates and the recombinant ADHs supported the observed stereoselectivity and confirmed the absolute configuration of the alcohol products. In summary, the present study provides efficient access to valuable drug-like enantiopure alcohols suitable for inclusion in chiral fragment-based drug discovery (FBDD) libraries. Key points Microbial whole-cell ketoreductases were investigated in bioreductions. Optimization of the reaction conditions greatly increased conversions. A docking study confirmed the enantiotopic selectivity of the ADHs.