Force fields matter in <scp>DNA</scp> pol η$$ \eta $$ mechanistic analysis

DNA polymerase <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML"> <mml:semantics><mml:mrow><mml:mi>η</mml:mi></mml:mrow> <mml:annotation>$$ \eta $$</mml:annotation></mml:semantics> </mml:math> is of major interest due to debates regarding the role of a third non-canonical magnesium ion in catalysis. By examining molecular simulations motivated by this mechanistic question and the role of the S113A mutation, we investigate how modern classical force fields compare for treating protein-DNA complexes and divalent metal ions in enzyme active sites. We find that while the CHARMM36m protein force field is robust, the nucleic acid components are unstable and too flexible, leading to the active site conformation in the reactant state not being well maintained. The addition of the third magnesium ion reduces but does not eliminate these issues, and the observed trends do not match experimental data and the magnesium ions bind too strongly. The AMBER19 force field leads to more stable nucleic acid components, but the standard 12-6 magnesium ion parameters do not reproduce experimental results on magnesium binding either. After comparing these models with the 12-6-4 ion models from OL15, General Amber force field GAFF-2, and re-parametrized m12-6-4 set, we find that the m12-6-4 model best aligns with experimental evidence and semi-empirical QM(DFTB3)/MM simulations. With these parameters, we conclude that the third magnesium ion binds transiently to the reactant state and strongly to the product state, stabilizing key active site structural features in both. The S113A mutation predominately disrupts the local water network of the active site.