Identification of the Catalytic Residues in the Cyclase Domain of the Class IV Lanthipeptide Synthetase SgbL

Lanthipeptides belong to the family of ribosomally synthesized and post‐translationally modified peptides (RiPPs) and are subdivided into different classes based on their processing enzymes. The three‐domain class IV lanthipeptide synthetases (LanL enzymes) consist of N‐terminal lyase, central kinase, and C‐terminal cyclase domains. While the catalytic residues of the kinase domains (mediating ATP‐dependent Ser/Thr phosphorylations) and the lyase domains (carrying out subsequent phosphoserine/phosphothreonine (pSer/pThr) eliminations to yield dehydroalanine/dehydrobutyrine (Dha/Dhb) residues) have been characterized previously, such studies are missing for LanL cyclase domains. To close this gap of knowledge, this study reports on the identification and validation of the catalytic residues in the cyclase domain of the class IV lanthipeptide synthetase SgbL, which facilitate the nucleophilic attacks by Cys thiols on Dha/Dhb residues for the formation of β‐thioether crosslinks.