Easy Solutions for Difficult Targets: Thermal Unfolding of GPCRs

Label-free Thermal Unfolding Assay of G Protein-Coupled Receptors for Compound Screening and Buffer Composition Optimization Using nanoDSF

Introduction

In this work, we apply a thermal unfolding-based assay using low volume differential intrinsic tryptophan scanning fluorimetry (nanoDSF) to study the stabilizing effects of ligands on G protein-coupled receptors (GPCRs). GPCRs are the fourth largest superfamily in the human genome and make up the largest class of drug targets. We focused on the human adenosine A2A receptor (A2AR), which differentially binds natural (adenosine and caffeine) and synthetic ligands to mediate a variety of physiological and pharmacological responses. We used several well-characterized ligands to perform unfolding experiments using nanoDSF. The resulting ΔTm shift values correlated well with data obtained using traditional ligand binding assays.