The ligand binding of G Protein-Coupled Receptors (GPCRs) is well studied, with a diverse literature examining binding events using computational and experimental methods to determine binding characteristics. These models are typically built through mass-action kinetics using variations of a single-site binding model for receptor-ligand interactions. With a few exceptions, computational models are built by extrapolating from previously reported parameters, to simulate the receptor-ligand complex formation from equilibrated ligand-binding data. In other cases, analytical solutions to the mass-action model have been used under restrictive experimental conditions to directly compute the system parameters.Based on experimental observations of both very fast and very slow kinetics via fluorescence anisotropy, we have implemented an analytical method that allows the direct computation of the kinetic parameters of a GPCR-ligand binding system, without the constraints imposed using analytical solutions. In addition, use of unlabeled ligand in competition with fluorescent ligand enables the calculation of the binding parameters for unlabeled ligand, making this an attractive option for screening of ligand properties as well as rapid screening of receptor properties. This promises an effective and rapid approach to drug screening applications.

Kinetic properties of G Protein Coupled Receptor ligand binding by anisotropic fluorimetry and computational modeling

247th National Meeting (2014)

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