Characterization by ENDOR Spectroscopy of the Iron–Alkyl Bond in a Synthetic Counterpart of Organometallic Intermediates in Radical SAM Enzymes

Members of the radical S-adenosyl-l-methionine (SAM) enzyme superfamily initiate a broad spectrum of radical transformations through reductive cleavage of SAM by a [4Fe–4S]1+ cluster it coordinates to generate the reactive 5′-deoxyadenosyl radical (5′-dAdo•). However, 5′-dAdo• is not directly liberated for reaction and instead binds to the unique Fe of the cluster to create the catalytically competent S = 1/2 organometallic intermediate Ω. An alternative mode of reductive SAM cleavage, especially seen photochemically, instead liberates CH3•, which forms the analogous S = 1/2 organometallic intermediate with an Fe–CH3 bond, ΩM. The presence of a covalent Fe–C bond in both structures was established by the ENDOR observation of 13C and 1H hyperfine couplings to the alkyl groups that show isotropic components indicative of Fe–C bond covalency. The synthetic [Fe4S4]3+–CH3 cluster, M-CH3, is a crystallographically characterized analogue to ΩM that exhibits the same [Fe4S4]3+ cluster state as Ω and ΩM, and thus an analysis of its spectroscopic propertiesand comparison with those of Ω and ΩMcan be grounded in its crystal structure. We report cryogenic (2 K) EPR and 13C/1/2H ENDOR measurements on isotopically labeled M-CH3. At low temperatures, the complex exhibits EPR spectra from two distinct conformers/subpopulations. ENDOR shows that at 2 K, one contains a static methyl, but in the other, the methyl undergoes rapid tunneling/hopping rotation about the Fe–CH3 bond. This generates an averaged hyperfine coupling tensor whose analysis requires an extended treatment of rotational averaging. The methyl group 13C/1/2H hyperfine couplings are compared with the corresponding values for Ω and ΩM