The radical S-adenosyl methionine (SAM) enzyme superfamily has widespread roles in hydrogen atom abstraction reactions of crucial biological importance. In these enzymes, reductive cleavage of SAM bound to a [4Fe-4S]1+ cluster generates the 5′-deoxyadenosyl radical (5′-dAdo•) which ultimately abstracts an H atom from the substrate. However, overwhelming experimental evidence has surprisingly revealed an obligatory organometallic intermediate Ω exhibiting an Fe-C5′-adenosyl bond, whose properties are the target of this theoretical investigation. We report a readily applied, two-configuration version of broken symmetry DFT, denoted 2C-DFT, designed to allow the accurate description of the hyperfine coupling constants and g-tensors of an alkyl group bound to a multimetallic iron–sulfur cluster. This approach has been validated by the excellent agreement of its results both with those of multiconfigurational complete active space self-consistent field computations for a series of model complexes and with the results from electron nuclear double-resonance/electron paramagnetic resonance spectroscopic studies for the crystallographically characterized complex, M–CH3, a [4Fe-4S] cluster with a Fe–CH3 bond. The likewise excellent agreement between spectroscopic results and 2C-DFT computations for Ω confirm its identity as an organometallic complex with a bond between an Fe of the [4Fe-4S] cluster and C5′ of the deoxyadenosyl moiety, as first proposed.

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-CH 3 , 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 propertiesand comparison with those of Ω and ΩMcan be grounded in its crystal structure. We report cryogenic (2 K) EPR and 13C/1/2H ENDOR measurements on isotopically labeled M-CH 3 . 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.