Radical Propagation via σ-Cleavage Mediates Radical SAM Catalyzed Sulfur-for-Oxygen Swapping Reaction during the Biosynthesis of Albomycin δ.

AbmM is a radical -adenosyl l-methionine (SAM) enzyme that catalyzes a radical initiated sulfur-for-oxygen swapping reaction, transforming the furanose ring of cytidine diphosphate (CDP) to a 4'-hydroxy-4'-thiofuranose product. While the function of AbmM has been demonstrated, the underlying mechanism regarding the formation of the radical intermediates during the reaction pathway remains to be fully established. To gain additional insight into this vital step in the biosynthesis of albomycin δ, 2'-deoxy-2'-methylidene CDP was synthesized as a mechanistic probe. Upon incubation with AbmM and dithionite, a C1' radical intermediate is generated from this mechanistic probe in the form of an allylic radical that can be trapped via oxidation to a sulfinate or a sulfenate versus reduction. Moreover, incubation of 2'-deoxy-2'-spirocyclopropryl CDP with AbmM also leads to a C1' radical intermediate that triggers opening of the cyclopropane ring. In this case, however, the resulting C7' terminal radical is not directly quenched but instead adds to the C5═C6 double bond of the cytosine base to form a new C7'-C6 bond. Taken together, these studies establish the intermediacy of a C1' radical species and thus suggest radical propagation from the C4' radical to the C1' radical through cleavage of the C1'-O bond prior to the sulfur insertion step during the AbmM-catalyzed reaction.