The radical S-adenosyl-L-methionine (SAM) enzyme superfamily is one of the largest known in Nature, with nearly 106 unique sequences spanning all kingdoms of life. Radical SAM reactions are remarkably diverse, and include difficult bond-breaking, bond-making, and complex rearrangements that are central to key metabolic pathways. Radical SAM catalysis is initiated by transfer of an electron from a reduced, site-differentiated [4Fe-4S]+ cluster to a SAM coordinated to the unique iron of the cluster, leading to homolytic cleavage of the S-C5′ bond of SAM and initiation of radical chemistry. The first observed intermediate is the organometallic Ω, in which C5′ of the SAM-derived deoxyadenosyl moiety is directly bound to the unique iron of the [4Fe-4S] cluster. The initial discovery of Ω was surprising, but Ω has since been observed in a wide range of radical SAM enzymes, and is found to be mechanistically central to radical initiation. The 5′-deoxyadenosyl radical (5′-dAdo●), long implicated in radical SAM reactions, is formed subsequent to Ω, consistent with a mechanism in which the Fe-C5′ bond of Ω undergoes homolysis to liberate the 5′-deoxyadenosyl radical for reaction with substrate. We have succeeded in trapping the 5′-dAdo● radical during substrate induced turnover conditions, and have shown progression of the reaction from 5′-dAdo● to a substrate-based product radical, demonstrating the catalytic competence of the 5′-deoxyadenosyl radical as a true intermediate in radical SAM catalysis. Together, our results provide a clear picture of the progression of steps during radical initiation in radical SAM reactions, with Ω undergoing homolytic Fe-C5′ bond cleavage to release a 5′-deoxyadenosyl radical, which then attacks substrate either by H-atom abstraction or adenosylation. This series of steps parallels the initiation of radical chemistry by adenosylcobalamin-dependent B12 enzymes, revealing the common use of organometallic chemistry to generate the central 5′-dAdo● radical species utilized by these two distinctly different classes of radical-based enzymes. The mechanistic picture of radical SAM catalysis provides important clues to how Nature harnesses, with exquisite regioselective and stereoselective control, the highly reactive 5′-deoxyadenosyl primary carbon radical for initiation of an incredible range of radical-based chemical transformations essential to life.

Mechanism of radical initiation in radical SAM enzymes

Where thousands of chemistry professionals meet to share ideas and advance scientific and technical knowledge. Spring 2023 abstracts, posters, and presentations will launch at the end of March.

ACS Spring 2023

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.