Figure 11. Aqueous ligand pK_a can influence overall protein-ligand binding affinity.

A When only the minor aqueous protonation state contributes to protein-ligand complex formation, the overall binding free energy (ΔG_bind) needs to be calculated as the sum of binding affinity of the minor state and the protonation penalty of that state. B When multiple charge states contribute to complex formation, the overall free energy of binding includes a multiple protonation states correction (MPSC) term (ΔG_corr). MPSC is a function of pH, aqueous pK_a of the ligand, and the difference between the binding free energy of charged and neutral species $(\Delta G_{bind}^C-\Delta G_{bind}^N)$.