α-Ketoglutarate-Dependent Dioxygenase-Catalyzed Macrocyclization in RiPPs

α-Ketoglutarate-dependent dioxygenases (αKGDs) are versatile biocatalysts for oxidative reactions, yet their role in ribosomally synthesized and post-translationally modified peptide (RiPP) macrocyclization remains unexplored. In this study, we uncover the widespread co-occurrence of JmjC domain-containing αKGDs (JMJDs) with diverse RiPP subfamilies in biosynthetic gene clusters (BGCs) through bioinformatic analysis. Specifically, we identify a conserved group of class II lanthipeptide BGCs encoding a two-component JMJD pair. As a representative example, heterologous reconstitution of the cor BGC from Streptomyces corchorusii, encoding a class II lanthipeptide synthetase CorM and the αKGD pair CorB-CorD, yields the metal-binding lanthipeptide corsin, which features a methyllanthionine (MeLan) ring and a unique Cys(S)-Tyr(Cβ) cross-link. Functional studies reveal that CorM installs a MeLan ring in the precursor peptide CorA, while the CorB-CorD pair catalyzes oxidative Cys(S)-Tyr(Cβ) cross-linking. Mechanistic investigations demonstrate that CorD, a canonical JMJD, catalyzes the Tyr7 Cβ-H abstraction and generates a para-quinone methide (pQM) intermediate, which undergoes Michael addition with Cys3. CorB, an inactive JMJD homologue, is essential for catalysis, unveiling a unique two-component αKGD system. This work expands the RiPP enzymatic toolkit by introducing αKGDs as radical catalysts for macrocyclization and advances our understanding of the diverse enzymatic strategies driving RiPP structural diversification.