G-protein coupled receptors (GPCRs) are the largest group of integral membrane proteins, and currently more than 30% of drugs target this superfamily. α1-Adrenoreceptors belong to GPCRs which are involved in many physiological processes in the brain and heart and consequently are linked to diseases such as heart failure and benign prostatic hyperplasia. However, biochemical, and structural characterization, as well as drug discovery, is often limited by the inherently unstable nature of membrane proteins. Currently, membrane protein including GPCRs requires to be purified in detergent micelles to enable structure determination through X-ray crystallography, Cryo-EM, and NMR. These artificial environments can impact the natural dynamics of the receptors and also interfere with a wide range of downstream experiments used to characterize structure and function such as NMR, Mass spectroscopy, ELISA, and pharmacological studies.
As a robust tool, Nanometer-scale discoidal phospholipid bilayers including Nanodiscs, Saposin nanoparticles, and Peptidiscs have been introduced to tackle this challenge, providing a more native-like environment and improving sample longevity. They typically contain an embedded membrane protein in a phospholipid bilayer which is encapsulated by a multisubunit protein belt. We are using Saposin nanoparticles to biophysically characterize the α1A-adrenoceptor. Experiments include elucidating ligand-dependent conformational changes of α1A-adrenoceptor by NMR using 13CH3-Met and 19F-Cys labeling of the receptor. The completion of this project will give fundamental insights into the molecular underpinnings of ligand binding and receptor activation.