The redox biosynthesis system has important applications in green biomanufacturing of chiral compounds. Formate dehydrogenase (FDH) catalyzes the oxidation of formate into carbon dioxide, which is associated with the reduction of NAD(P)⁺ into NAD(P)H. Due to this property, FDH is used as a crucial enzyme in the redox biosynthesis system for cofactor regeneration. Nevertheless, the application of natural FDH in industrial production is hampered by low catalytic efficiency, poor stability, and inefficient coenzyme utilization. This review summarized the structural characteristics and catalytic mechanism of FDH, as well as the advances in protein engineering of FDHs toward improved enzyme activity, catalytic efficiency, stability and coenzyme preference. The applications of using FDH as a coenzyme regeneration system for green biomanufacturing of chiral compounds were summarized.