All starting structures were then solvated in a 24 Å spherical droplet of TIP3P water molecules, centered on the C1 atom of bound DHAP or GAP (see ref. for atom numbering), described using the Surface Constrained All Atom Solvent (SCAAS) approach. All residues within the inner 85% of this sphere were fully mobile, whereas those in the outer 15% were subjected to a 10 kcal mol -1 Å -2 harmonic positional restraint to restrain them to their original crystallographic positions, and all residues outside the droplet were essentially fixed to their crystallographic positions using a 200 kcal mol -1 Å -2 restraint, as described in previous work. All residues within the mobile region were kept in their expected ionization states at physiological pH, assigned based on a combination of analysis using PROPKA 3.1 and visual inspection, and residues outside the mobile region were kept in their neutral form, as is standard practice in such simulations in order to avoid system instabilities introduced due to the presence of charges outside the explicit simulation sphere. For a full list of ionized residues and histidine protonation states in our simulations, see Table .