The LacI-Type Transcriptional Activator NCgl2689 Directly Regulates an Operon Involved in l-Cysteine Synthesis by the Reverse Transsulfuration Pathway in Corynebacterium glutamicum

l-Cysteine is widely used in food, pharmaceutical, animal feed, and cosmetic industries. Microbial fermentation is an economically effective method for producing l-cysteine. However, research on the synthesis and regulation mechanism of l-cysteine through the reverse transsulfuration pathway in important amino acid producing cell factory Corynebacterium glutamicum is still limited. In this study, we demonstrated that NCgl2688 had cystathionine γ-lyase (CGL) activity (2.98 × 10² M^-1 s^-1), which played an important role in the conversion of cystathionine to l-cysteine by the reverse transsulfuration pathway. In the ΔNCgl2689 strain, the mRNAs of NCgl2689 and the l-cysteine biosynthetic gene NCgl2688 were significantly reduced and the activity of NCgl2688 was relatively low. Moreover, the expression of the NCgl2688 or NCgl2689 gene was significantly upregulated in response to H₂O₂ or l-methionine. The ability of NCgl2689 to bind to the promoter region of the NCgl2689 and NCgl2688 genes was confirmed through EMSA. Furthermore, the growth of the ΔNCgl2689 strain was significantly inhibited by antibiotics, alkylating agents, or oxidants. This work reveals that NCgl2689 is the first regulator of CGL, converting cystathionine into l-cysteine in Corynebacteriales, and provides new insights into the regulatory mechanisms involved in the stress resistance and l-cysteine biosynthesis through the reverse transsulfuration pathway.