Insulin dimer dissociation in aqueous solution: A computational study of free energy landscape and evolving microscopic structure along the reaction pathway.

The dissociation of an insulin dimer to two monomers is an important life process. Although the monomer is the biologically active form of the hormone, it is stored in the β-cells of the pancreas in the hexameric form. The latter, when the need comes, dissociates to dimers and the dimers in turn to monomers to maintain the endogenous delivery of the hormone. In order to understand insulin dimer dissociation at a molecular level, we perform biased molecular dynamics simulations (parallel tempering metadynamics in the well-tempered ensemble) of the dissociation of the insulin dimer in water using two order parameters and an all-atom model of the protein in explicit water. The two order parameters selected (after appropriate studies) are the distance (RMM) between the center of mass of two monomers and the number of contacts (NMM) among the backbone-Cα atoms of the two monomers. We calculated the free energy landscape as a function of these two order parameters and determined the minimum free energy pathway of dissociation. We find that the pathway involves multiple minima and multiple barriers. In the initial stage of dissociation, the distance between the monomers does not change significantly but the NMM decreases rapidly. In the latter stage of separation, the opposite occurs, that is, the distance RMM increases at nearly a constant low value of NMM. The configurations of the two monomeric proteins so formed are found to be a bit different due to the entropic reasons. Water is seen to play a key role in the dissociation process stabilizing the intermediates along the reaction path. Our study reveals interesting molecular details during the dissociation, such as the variation in the structural and relative orientational arrangement of the amino acid residues along the minimum energy path. The conformational changes of monomeric insulin in the stable dimer and in the intermediate states during dimer dissociation have been studied in detail.