Apparent thermodynamic parameters of ligand binding to the cloned rat μ-opioid receptor

The apparent thermodynamic parameters of binding of ten ligands to the cloned rat μ-opioid receptor stably expressed in Chinese hamster ovary (CHO) cells were investigated. For every ligand, the K d or K i values at 0°C, 12°C, 25°C and 37°C were determined, a van't Hoff plot was generated and Δ H°′, Δ S°′ and − TΔ S°′ and Δ G°′ were calculated. Changes in free energy (Δ G°′) ranged from −10.35 to −15.65 kcal/mol. The binding of sufentanil, ohmefentanyl, diprenorphine and d-Phe-Cys-Tyr- d-Trp-Arg-Thr-penicillamine-Thr-NH 2 (CTAP) was endothermic (Δ H°′>0) and driven by an increase in entropy (− TΔ S°′=−13.08 to −18.57 kcal/mol). The binding of naltrexone was exothermic (Δ H°′=−12.56 kcal/mol) and essentially enthalpy-driven. The binding of morphine, methadone, pentazocine, [ d-Ala 2, NMePhe 4, Gly-ol]enkephalin (DAMGO) and Tyr-Pro-NMePhe- d-Pro-NH 2 (PL017) was exothermic (Δ H°′=−3.53 to −9.95 kcal/mol) and occurred with an increase in entropy (− TΔ S°′=−2.48 to −7.92 kcal/mol). Plots of enthalpy versus entropy and enthalpy versus free energy were linear, although enthalpy–entropy compensation was not evident. The entropy changes were not correlated with apparent lipophilicity of the compounds. These results suggest that: (1) opioid ligands bind to the μ receptor by specific mechanisms, unrelated to lipid solubility; (2) the mechanism of binding is not universally different for peptide and non-peptide ligands; (3) the nature of binding does not a priori determine intrinsic activity. The results reveal a novel differentiation of opioid ligands into two groups (group 1: ohmefentanyl, sufentanil, diprenorphine, CTAP and PL017; group 2: naltrexone, morphine, methadone, DAMGO, pentazocine), based on two distinct relationships between enthalpy versus free energy of binding, the details of which are yet to be elucidated.