| dc.description | Although it is recognized that electrostatic potential roughly shapes the energetic landscape of macromolecules, the scale and the number of interacting entities in these systems hinders the quantitative appreciation of this contribution. The biophysics of two electrostatically-sensitive equilibria in cytochrome c has been studied using protein engineering. First, a pH-induced ligand exchange known as the alkaline transition was investigated using an (alternative) linear regression of titration curves. The alkaline transition proved to be a 2 step equilibrium that is led by the deprotonation of the high-pH ligands and another ionization, thought to trigger this cooperative change. Second, the thermodynamics of oxido-reduction of cytochrome c was investigated by point-charge probing and by engineering the internal polarity of the protein matrix. From the discrimination between enthalpic and entropic contribution to the equilibrium, it appears that the dielectric susceptibility is constant within the protein matrix. However, an entropic factor thought to be conformational polarisability dominates the dielectric effect. This explains why no predictable dielectric constant can be derived directly from Coulom's law. It also appears that dipolar and conformational susceptibilities combine in a form of homeostasis that preserves the unction of the cytochrome. This functional cooperativity is proposed to confer the robustness and flexibility necessary for the protein to survive (near-)random mutations throughout evolution. | en_US |
