Research Progress on the Regeneration of the Coenzyme of Flavin-Dependent Halogenases

Considerable and increasing attention is being directed toward the application of halogenases for the biocatalytic functionalization of C-H bonds. Biocatalytic approaches offer distinct advantages over conventional chemical methods, including excellent catalyst-controlled selectivity and the avoidance of protecting groups and hazardous reagents. Ongoing discovery of natural halogenation pathways has led to the identification and characterization of a growing number of halogenases. Flavin-dependent halogenases (FDHs) have emerged as a major research focus, driven by their utility in the biohalogenation of aromatic scaffolds. These enzymes exhibit high regioselectivity and operate under environmentally benign conditions, making them attractive catalysts for the sustainable synthesis of halogenated compounds. Nevertheless, the activity of FDHs depends on reduced flavin adenine dinucleotide (FADH₂), and the efficient regeneration of this cofactor remains a critical bottleneck hindering industrial implementation. This review comprehensively summarizes recent advances in cofactor regeneration strategies for FDH. Herein, we discuss the mechanisms, advantages, and applicability of various regeneration systems, including traditional multienzyme cascades and emerging single-component self-sufficient architectures, finally concluding with an outlook on future research directions.