The use of microcalorimetry to measure thermodynamic parameters of the binding of ligands to insulin.

Flow microcalorimetry was used to measure the free energies, enthalpies, and entropies of interactions between the hormone insulin and small ligand molecules or ions. Measurable amounts of heat were obtained for binding of four phenolic preservative molecules--phenol, meta-cresol, resorcinol, and methylparaben--to both two-zinc and zinc-free insulin and for binding of zinc ions to zinc-free insulin. All of the reactions were spontaneous, but the phenolic binding was driven by enthalpy, while that of zinc was entropy-driven. A combination of van der Waals interactions, hydrophobic effects, and protein conformational changes appeared to be involved in binding of the phenolic ligands. Zinc ions displayed two types of binding to insulin, both involving ion-dipole interactions.