New Polymer Belts for Lipid-Bilayer Nanodiscs to Study the Molecular Basis of GPCR Signalling

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Membrane proteins play fundamental roles in cellular communication and material transfer and account for about half of all drug targets. However, their extraction, purification, and in vitro investigation are very challenging. Current methods used for isolating membrane proteins often depend on aggressive chemistry and result in protein denaturation. This is also true for many pharmacological target proteins, whose poor stability upon removal from the native membrane environment seriously impairs drug discovery. Among new chemically inspired strategies, nano-discs assembled from lipids and styrene/maleic acid (SMA) copolymers have received great attention. These polymers can recruit membrane proteins and lipids directly from membranes into nanoscale lipid-bilayer patches. Yet, this promising technology is currently limited by alterations in the conformational dynamics of embedded membrane proteins and lipids. Further limitations are the presence of UV-absorbing aromatic rings and the high charge density due to the polymer’s carboxyl groups.NanoBelt will develop new polymers that combine good solubilisation yields with low UV absorption, negligible charge density, and maintenance of the function and conformational dynamics of proteins. Polymers are prepared by Partner 1 (IBMM—Avignon University) by post-functionalisation of existing polymers or de novo synthesis from functionalised monomers. Partner 2 (TU Kaiserslautern) determines the potency of the new polymers to form nanodiscs and to extract membrane proteins to select optimal polymers for sensitive G protein-coupled receptors (GPCRs). The ghrelin receptor (GHSR) is used as a prototypical class A GPCR by Partner 3 (IBMM—CNRS). To this end, the recombinant receptor is assembled into the new nanodiscs for determination of its pharmacological properties. Partner 4 (Leipzig University) uses the corticotropin releasing factor receptor (CRF1R) as a model of class B GPCRs. Using new FRET sensors, the conformational dynamics of these challenging drug targets will be dissected in the native-like but controlled environment of new polymer-based nanodiscs. Owing to a systematic approach of validation based on the complementary skills of Partners 2, 3, and 4, NanoBelt will provide structure–activity relationships to enable the rational design of improved polymers synthesised by Partner 1.The ultimate goal consists in identifying and establishing new polymers that extract membrane proteins with their surrounding lipids directly from cellular membranes in a mild yet efficient manner to form polymer-encapsulated lipid-bilayer nanodiscs that retain the native structures and functions of the extracted proteins.