Critical evaluation of two neutron structures of Mn superoxide dismutase with quantum refinement

In 2021, neutron structures of oxidised and reduced human manganese superoxide dismutase were published, suggesting several unexpected features, including deprotonated glutamate, tyrosine and histidine residues and a reduced Mn ion with two hydroxide ligands. We have used quantum refinement to evaluate whether alternative interpretations of the structures are possible, comparing many structural models with different possible protonation states or other structural interpretations. Quantum refinement is standard crystallographic refinement in which the empirical restraints, which are employed to ensure that the structure makes chemical sense and gives reasonable bond lengths and angles, are replaced by quantum mechanical calculations for a small but interesting part of the structure. We show that in all cases, there are more chemically reasonable interpretations of the structures, not involving any deprotonated residues, which give slightly improved structures in terms of real-space Z scores based on the difference maps and strain energies. Weak nuclear densities do not necessarily indicate that an atom is not present; instead it may indicate that the atom may have several conformations or shows extensive dynamics. For the Tyr-34 residue, our results indicate that the phenolic hydroxide H atom should preferably be within the aromatic ring plane (by ~20 kJ/mol), but with two possible conformations. Moreover, an Mn-bound water molecule can readily accept a hydrogen bond from the nearby Gln-143 residue.