Effects of histidine protonation and phosphorylation on histidine-containing phosphocarrier protein structure, dynamics, and physicochemical properties

Previous structural studies of the histidine-containing phosphocarrier protein (HPr) have shown that active site residue His15 can adopt two distinct conformations which were termed OPEN and CLOSED. Using molecular dynamics simulations and protonation probability calculations, we were able to show that these two conformations correspond to different protonation forms of the histidine ring. The CLOSED-to-OPEN transition requires His15 to adopt a conformation with higher energy, which is compensated by the favorable energetic consequences of protonation. Calculations of the conformational energy of His15 show that HPr exists mainly in the CLOSED form at pH 7. The very low apparent pKa value (3.2−4.5) of the CLOSED conformation and the fact that the imidazole ring of residue 15 is primarily unprotonated at Nδ1 at neutral pH ensure that His15 is ideally primed to be specifically phosphorylated at Nδ1. In contrast to unphosphorylated HPr, the phosphorylated form exhibits no conformational transitions, and the ...