Proteins are polyelectrolytes with acidic and basic amino acids Asp, Glu, Arg, Lys, and His, making up ≈25% of the residues. The protonation state of residues, cofactors, and ligands defines a "protonation microstate". In an ensemble of proteins some residues will be ionized and others neutral, leading to a mixture of protonation microstates rather than in a single one as is often assumed. The microstate distribution changes with pH. The protein environment also modifies residue proton affinity so microstate distributions change in different reaction intermediates or as ligands are bound. Particular protonation microstates may be required for function, while others exist simply because there are many states with similar energy. Here, the protonation microstates generated in Monte Carlo sampling in MCCE are characterized in HEW lysozyme as a function of pH and bacterial photosynthetic reaction centers (RCs) in different reaction intermediates. The lowest energy and highest probability microstates are compared. The Δ, Δ, and Δ between the four protonation states of Glu35 and Asp52 in lysozyme are shown to be calculated with reasonable precision. At pH 7 the lysozyme charge ranges from 6 to 10, with 24 accepted protonation microstates, while RCs have ≈50,000. A weighted Pearson correlation analysis shows coupling between residue protonation states in RCs and how they change when the quinone in the Q site is reduced. Protonation microstates can be used to define input MD parameters and provide insight into the motion of protons coupled to reactions.