Shedding light on reactive sulfur species coordination to metmyoglobin by QM–MM TD-DFT simulations

Understanding how sulfur-containing ligands modulate the structure, electronic configuration, and spectroscopy of ferric heme proteins is essential for interpreting their reactivity and assigning experimentally observed intermediates. Here, we investigated the spectroscopic properties of a series of ferric myoglobin species, MbFe(III)-X, being X = HS-, S2-, H2S2, HS2-, S22-, and OH-. We combined QM-MM molecular dynamics (treating the active site and the coordinated ligands as the QM region, with the rest of the protein and solvent treated classically) with the nuclear ensemble approach, in which we sampled configurations from the MD simulations and obtained the electronic spectra of the species using linear-response TD-DFT at the B3LYP/def2-TZVP level of theory. The simulated spectra reproduce the major experimental trends, providing strong support for assigning the sulfide-bound intermediate to MbFe(III)-HS-, explaining the near indistinguishability of MbFe(III)-HS2- and MbFe(III)-S22- spectra, and rationalizing the similarities between sulfide- and disulfide-derivatives absorption profiles. Taken together, our results demonstrate the importance of explicitly sampling nuclear configurations to account for dynamic fluctuations in the active sites of complex metalloproteins when modeling their electronic spectra. By incorporating this configurational heterogeneity, the resulting spectral predictions achieved a high level of reliability and showed close agreement with the experiments.