Determination of equilibrium dissociation constants in fluorescence polarization.
A simple mathematical model (the FP K(d) model) is used to generate the dissociation equilibrium constant (K(d)) for G protein-coupled receptor-ligand binding measured using fluorescence polarization (FP) saturation curve analysis. The model generates data that may be analyzed by the method of Scatchard. The validity of the FP K(d) model is proven in six model systems in which the modeled K(d) values are within a factor of 5 of inhibitory equilibrium constant values obtained from radioligand competition assays.
Prystay
L
L
PerkinElmer Life Sciences, Montreal, Quebec, Canada.
Gosselin
M
M
Banks
P
P
D001667:Binding, Competitive; D019793:Fluorescein; D005454:Fluorescence Polarization / Q000379:methods*; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008956:Models, Chemical; D011485:Protein Binding; D018043:Receptors, Corticotropin / Q000378:metabolism; D044101:Receptors, Melanocortin; D015203:Reproducibility of Results; D013816:Thermodynamics; D000521:alpha-MSH / Q000378:metabolism
D008024:Ligands; D018043:Receptors, Corticotropin; D044101:Receptors, Melanocortin; C086971:melanocortin 5 receptor; D000521:alpha-MSH; D020962:Heterotrimeric GTP-Binding Proteins; D019793:Fluorescein