Binding kinetics of five drugs to beta2-adrenoceptor using peak profiling method and nonlinear chromatography

Investigations of drug-protein interactions have advanced our knowledge of ways to design more rational drugs. In addition to extensive thermodynamic studies, ongoing works are needed to enhance the exploration of drug-protein binding kinetics. In this work, the beta2-adrenoceptor (β2-AR) was immobilized on N, N'-carbonyldiimidazole activated amino polystyrene microspheres to prepare an affinity column (4.6 mm × 5.0 cm, 8 μm). The β2-AR column was utilized to determine the binding kinetics of five drugs to the receptor. Introducing peak profiling method into this receptor chromatographic analysis, we determined the dissociation rate constants (kd) of salbutamol, terbutaline, methoxyphenamine, isoprenaline hydrochloride and ephedrine hydrochloride to β2-AR to be 15 (±1), 22 (±1), 3.3 (±0.2), 2.3 (±0.2) and 2.1 (±0.1) s−1, respectively. The employment of nonlinear chromatography (NLC) in this case exhibited the same rank order of kd values for the five drugs bound to β2-AR. We confirmed that both the peak profiling method and NLC were capable of routine measurement of receptor-drug binding kinetics. Compared with the peak profiling method, NLC was advantageous in the simultaneous assessment of the kinetic and apparent thermodynamic parameters. It will become a powerful method for high throughput drug-receptor interaction analysis.