When large QM regions have been used in multi-scale quantum-mechanical-molecularmechanical (QM/MM) simulations of enzyme catalysis, distinct nuclear and charge dynamics have been observed in comparison to those using small QM regions. This is consistent with observations that much of the protein must be studied quantum mechanically to obtain asymptotic convergence of QM properties such as the favorability of proton or charge transfer , electric fields , excitation energies , bond critical points and partial charges . Nevertheless, conclusions from QM/MM studies of charge fluctuations are potentially limited by the significant effects of the boundary treatment and embedding method on the charge distribution in the QM region. Although some QM-derived effects can be captured using polarizable force field modeling , charge transfer and dynamical formation of charge-assisted hydrogen bonds remain outside the scope of those methods.