Abstract
\nDNA polymerase \u03b7 is of major interest due to debates regarding the role of a
\nthird 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
\nprotein-DNA complexes and divalent metal ions in enzyme active sites. We
\nfind that while the CHARMM36m protein force field is robust, the nucleic acid
\ncomponents are unstable and too flexible, leading to the active site conformation in the reactant state not being well maintained. The addition of the third
\nmagnesium ion reduces but does not eliminate these issues, and the observed
\ntrends do not match experimental data and the magnesium ions bind too
\nstrongly. The AMBER19 force field leads to more stable nucleic acid components, but the standard 12-6 magnesium ion parameters do not reproduce
\nexperimental results on magnesium binding either. After comparing these
\nmodels with the 12-6-4 ion models from OL15, General Amber force field
\nGAFF-2, and re-parametrized m12-6-4 set, we find that the m12-6-4 model
\nbest aligns with experimental evidence and semi-empirical QM(DFTB3)\/MM
\nsimulations. With these parameters, we conclude that the third magnesium ion
\nbinds 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.<\/p>\n