Advances in Cystathionine β-Lyase-Catalyzed Synthesis of Thiosulfinates: Enzyme Resource Mining, Catalytic Mechanism, and Structure-Guided Molecular Engineering

Abstract Thiosulfinates, represented by diallyl thiosulfinate (allicin), are the material basis of the characteristic flavor and bioactivity of Allium plants. They show broad-spectrum antibacterial, antibiofilm, antiviral, and antitumor activities. However, their extreme chemical instability challenges conventional plant extraction and chemical synthesis. In situ enzymatic synthesis, in which alliinase (l-cysteine sulfoxide lyase, CSL) generates thiosulfinates on demand from S-alk(en)yl-l-cysteine sulfoxide substrates, offers a new solution. Plant-derived CSLs are limited by difficult extraction and low heterologous expression efficiency, whereas bacterial cystathionine β-lyase (CBL), which is easily expressed at low cost, has emerged as a novel catalyst for thiosulfinate biosynthesis. This review summarizes thiosulfinate chemistry and biosynthesis, the structure and pyridoxal phosphate (PLP)-dependent catalysis of CBL, and the discovery of CBL’s CSL activity. Aiming at future food, agricultural, and pharmaceutical applications, we further analyze CBL’s bottlenecks in substrate stereoselectivity and pH preference, highlighting enzyme resource mining and molecular engineering strategies.