A biophysical basis of ligand-induced activation of excitable membranes and associated enzymes. A thermodynamic study using acetylcholinesterase as a model receptor

A biophysical analysis of current models for the molecular mechanism of ligand activation of receptors and regulatory enzymes is offered. Acceptance of the working hypothesis that the resting state P r of the receptor is formally in thermodynamic equilibrium with an energized state P e allows the prediction that the role of activator ligands should be to modulate that physical variable which controls the relative ground-state conformational energies of the two states. It is pointed out that for a majority of ligands nonexclusive binding is probably operative. The use of acetylcholinesterase (AChE) as a model receptor led to the finding that for some 30 ligands of the tetramethylammonium ion (TMA) series, the parameter ΔF binding is constant at −4.2 ± 0.6 kcal/mole, while the thermodynamic parameters ΔH and ΔS undergo large but compensated fluctuations. The additivity of ΔH and ΔS gives rise to an isoequilibrium relationship of slope β = 288 °K in agreement with an interaction mechanism where a ligand-induced release of "ice-like" water from the binding cleft is operative. Previous criticisms of the physical significance of isoequilibrium relations are shown to be largely inapplicable to the problem under consideration. The interaction mechanism is in accordance with the equation[Formula: see text]where ΔF = ΔH int = −4.15 kcal/mole (the ion–ion interaction), TΔS int = 0, and ΔH ext = ΔS ext = variable endothermic release of "ice-like" water molecules. It is shown that the TMA ligands strongly modulate the ΔH ext –ΔS ext couple. Since the release of peptide-bound water molecules is tantamount to a conformational change, it is concluded that the ligands stabilize in varying degrees of efficiency a predetermined conformational state (P e ) of the protein. The conclusion is reached that the ligand-induced perturbation theory of drug action and the induced-fit theory of enzyme action are valid at least in the physical sense.The parameters ΔH ext and ΔS ext are shown to be additive functions of the molar volume, the intrinsic flexibility, and the physical properties of the ligands. Large anomalous effects of structure and optical isomerism on ΔH ext and ΔS ext appear to correlate in some manner with high pharmacological potency. It is concluded that a ligand-induced modulation of the physical variable of ice melting from the binding cleft may control the position of the conformational equilibrium between the resting state P r and the energized state P e of the receptors.