Design of a self-sufficient hydride-shuttling cascade for concurrent bioproduction of 7,12-dioxolithocholate and l-tert-leucine

Abstract

Oxidoreductase-mediated biotransformation often requires consumption of a secondary sacrificial co-substrate and an additional auxiliary enzyme to drive the cofactor regeneration, which results in generation of unwanted by-product. Herein, we report a highly atom-economic self-sufficient hydride-shuttling cascade to concurrently obtain two pharmaceutically important building blocks (7,12-dioxo-lithocholic acid and L-tert-leucine) in which oxidation of cholic acid (CA) and reductive amination of trimethylpyruvic acid were integrated for redox self-recycling. In this cascade, the cofactor acts as a hydride shuttle that interconnects the two synthetically relevant reactions at the cost of only inorganic ammonium as the sacrificial agent and generates water as the greenest by-product. The preparative biotransformation using a whole-cell biocatalyst in the absence of any exogenous cofactor displayed a space–time yield of 768 g L⁻¹ d⁻¹ and a total turnover number (TTN) of 20 363 for NAD⁺ recycling. This represents the highest cofactor TTN reported to date for the bio-oxidation of CA, indicating the great potential of this cofactor and redox self-sufficient bioprocess for cost-effective and sustainable biomanufacturing of high-value-added products.