Polymer-Based Nanodiscs for Studying Structure and Dynamics of G-Protein-Coupled Receptors

Abstract

G-protein-coupled receptors (GPCRs) constitute the largest family of membrane proteins and are most important drug targets. Understanding of their inner workings is only in the beginning, because as integral membrane proteins GPCRs require the use of detergents to solubilize them into an aqueous environment for purification and subsequent biophysical and structural studies. Detergent micelles, however, are limited in their ability to mimic lipid bilayers. Recently, a promising approach was introduced that overcomes these detergent-related drawbacks: poly(styrene-co-maleic acid), a copolymer synthesized from styrene and maleic anhydride, has been shown to solubilize membrane proteins from various membranes without the need of detergents. In the resulting nanometer-sized nanodiscs the polymer wraps around the protein and its annular lipids. Here, we address the question to which extent polymer-based nanoparticles are able to stabilize a GPCR and how they influence protein activity and functionality.We compared activation of the GPCR and photoreceptor bovine rhodopsin and its equilibrium of metarhodopsin (Meta) photo-products for rhodopsin solubilized either in n-dodecyl-β-D-maltoside (DDM) detergent micelles or as polymer nanodiscs. While DDM favors active Meta II, the polymer-based environment favors the inactive Meta I and Meta III states, which might explain the lack of G-protein activation in the latter case. Protein function is re-established as soon as rhodopsin is transferred from the discs into detergent micelles or proteoliposomes. In contrast to DDM, the polymer belt also affects the binding of 11-cis-retinal ligand. Taken together, polymer nanodiscs are suited to extract GPCRs from native membranes in a detergent-free manner but appear to exert a rather rigid lipid environment prohibiting helical movements. The presented work is part of a study where we test various membrane-mimetic systems for studying structure and dynamics of GPCRs and other proteins related to Structural Neurobiology.