Integrated ESI-MS/EPR/computational characterization of the binding of metal species to proteins: vanadium drug-myoglobin application

An integrated experimental/computational strategy to study the binding modes of metal species to proteins is presented. With this multistep method based on the combined application of spectrometric (ESI-MS), spectroscopic (EPR) and computational (docking and QM) techniques, the interaction of VIVO2+ and four potential drugs VIVOL2 (L = 1,2-dimethyl-3-hydroxy-4(1H)-pyridinonate or dhp, L-mimosinate or mim, maltolate or ma, acetylacetonate or acac) with myoglobin (Mb) was characterized. ESI-MS allows the determination of the number of moieties (VOL+ or VOL2) bound to the protein, EPR helps distinguish the type of coordinating donors, and docking and full QM models allow the prediction of the specific residues involved in the V coordination as well as the 3D structure of the adducts. The results indicate that VIVO2+ ions bind to three different sites with the involvement of three residues of the polypeptide chain (His, Asp and Glu). In the systems with dhp and mim, mixed species {VOL2}n(Mb) with n = 2 (dhp) and 3-4 (mim) were formed with the equatorial coordination of one exposed His residue. With ma and acac, adducts with the general formula {VOL}n(Mb) with n = 2 were identified with the simultaneous binding of two residues (Glu, Asp or His) to two adjacent positions of the VOL+ moiety. This approach is generalizable and could be applied to other metal ions and proteins using - depending on the metal features - different spectroscopic techniques.