Green whole-cell biocatalysis: compartmentalized engineering of borneol dehydrogenase for stereoselective synthesis of (+)-borneol

Abstract

Stereoselective synthesis of stereochemically pure (+)-borneol—an important ingredient in traditional Chinese medicine and China-developed class I new drug Sanbexin®—remains challenging due to inherent limitations of chemical methods (harsh reagents, organic solvents, tedious chiral catalyst synthesis, poor stereocontrol, and a high global E factor) and existing biocatalysts (low efficiency, expensive purification, and insufficient stereocontrol). Herein, we report a green biocatalytic route leveraging a compartmentalized engineering strategy to reprogram Arabidopsis lyrata borneol dehydrogenase (AlBDH). By partitioning the enzyme into substrate-binding, loop, and catalytic microdomains, synergistic mutagenesis (focused rational iterative site-specific/iterative saturation/homology-guided) generated variant Mu3. This whole-cell biocatalyst mediates the reduction of (+)-camphor to (+)-borneol under mild aqueous conditions (25 °C, pH 6.2) using isopropanol for NADPH regeneration, achieving a 45% isolated yield and >99.5% de at 30 mM substrate loading on a gram scale. This work establishes a green, scalable platform for pharmaceutical-grade (+)-borneol production, advancing green chemistry in drug synthesis and traditional Chinese medicine modernization.