Dynamic residue ionization in a bacterial porin: Impact of ionic strength and membrane potential.

The protonation states of key residues play a critical role in regulating the function of membrane proteins. Also, protein channel characterization involves testing the effect of pH, membrane charge, ionic concentration, and applied potential as modulators of channel conductance, selectivity and gating. Using GROMACS Constant-pH Molecular Dynamics, here we investigate how ionic strength, externally applied voltage and structural conformation changes modulate the protonation state of the acidic residues of the OmpF channel. pKa shifts in response to increased KCl concentrations do not follow a simple pattern. This contrasts with earlier claims on the effect of ionic strength on the pKa of selected OmpF residues. Additionally, we find that applying an external voltage yields position-dependent small pKa shifts of opposite sign on the periplasmic and the extracellular side of the protein. This charge regulation coming from the external electric field adds to the well-known structural charge asymmetry of this channel. While pKa shifts involve minimal protein backbone conformation changes, there are some significant side chains rotations or displacements. Overall, these findings provide new insights into how different factors influence the protonation dynamics of OmpF. The results may have useful implications for other channels regulated by long-range electrostatic interactions.