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We investigate protonation as a possible route for triggering conformational change in proteins by focusing on the calmodulin (CaM) example. Two hundred nanosecond molecular dynamics (MD) simulations are performed on both the extended and compact forms of calcium loaded CaM. The stability of both structures is confirmed under prevailing conditions. Protonation of nine acidic residues with upshifted pK(a) values leads to a large conformational change in less than 100 ns. The structure attained is consistent with fluorescence resonance energy transfer experimental results as well as structures from an ensemble compatible with NMR data. Analysis of the MD trajectories summing up to one microsecond implies that the key events leading to the completion of the conformational change begins with an initial formation of a salt bridge between the N-lobe and the linker, followed by the bending of the C-lobe and the organization of a stabilizing hydrophobic patch between the lobes. We find that CaM utilizes its Ca2+ ions to harden/soften different regions so as to achieve various conformations. Thus, barrier crossing between extended and compact forms of CaM which is normally a rare event due to the repulsive electrostatic interactions between the two lobes is facilitated by protonation of high pK(a) residues. The results delineate how pH changes might be utilized in the cell to achieve different conformation-related functions.

Proteins are complex systems made up of many conformational sub-states which are mainly determined by the folded structure. External factors such as solvent type, temperature, pH and ionic strength play a very important role in the conformations sampled by proteins. Here we study the conformational multiplicity of calmodulin (CaM) which is a protein that plays an important role in calcium signaling pathways in the eukaryotic cells. CaM can bind to a variety of other proteins or small organic compounds, and mediates different physiological processes by activating various enzymes. Binding of calcium ions and proteins or small organic molecules to CaM induces large conformational changes that are distinct to each interacting partner. In particular, we discuss the effect of pH variation on the conformations of CaM. By using the pKa values of the charged residues as a basis to assign protonation states, the conformational changes induced in CaM by reducing the pH are studied by molecular dynamics simulations. Our current view suggests that at high pH, barrier crossing to the compact form is prevented by repulsive electrostatic interactions between the two lobes. At reduced pH, not only is barrier crossing facilitated by protonation of residues, but also conformations which are on average more compact are attained. The latter are in accordance with the fluorescence resonance energy transfer experiment results of other workers. The key events leading to the conformational change from the open to the compact conformation are (i) formation of a salt bridge between the N-lobe and the linker, stabilizing their relative motions, (ii) bending of the C-lobe towards the N-lobe, leading to a lowering of the interaction energy between the two-lobes, (iii) formation of a hydrophobic patch between the two lobes, further stabilizing the bent conformation by reducing the entropic cost of the compact form, (iv) sharing of a Ca+2 ion between the two lobes.