Effect of ethanol on insulin dimer dissociation.

Insulin-dimer dissociation is an essential biochemical process required for the activity of the hormone. We investigate this dissociation process at the molecular level in water and at the same time, in 5% and 10% water-ethanol mixtures. We compute the free energy surface of the protein dissociation processes by employing biased molecular dynamics simulation. In the presence of ethanol (EtOH), we observe a marked lowering in the free energy barrier of activation of dimer dissociation from that in the neat water, by as much as ∼50%, even in the 5% water-ethanol solution. In addition, ethanol is found to induce significant changes in the dissociation pathway. We extract the most probable conformations of the intermediate states along the minimum energy pathway in the case of all the three concentrations (EtOH mole fractions 0, 5, and 10). We explore the change in microscopic structures that occur in the presence of ethanol. Interestingly, we discover a stable intermediate state in the water-ethanol binary mixture where the centers of the monomers are separated by about 3 nm and the contact order parameter is close to zero. This intermediate is stabilized by the wetting of the interface between the two monomers by the preferential distribution of ethanol and water molecules. This wetting serves to reduce the free energy barrier significantly and thus results in an increase in the rate of dimer dissociation. We also analyze the solvation of the two monomers during the dissociation and both the proteins' departure from the native state configuration to obtain valuable insights into the dimer dissociation processes.