Capturing G protein-coupled receptors into native lipid-bilayer nanodiscs using new diisobutylene/maleic acid (DIBMA)...

Many membrane proteins, including G protein-coupled receptors (GPCRs), are susceptible to denaturation when extracted from their native membrane by detergents. Therefore, alternative methods have been developed, including amphiphilic copolymers that enable the direct extraction of functional membrane proteins along with their surrounding lipids. Among these amphiphilic copolymers, styrene/maleic acid (SMA) and diisobutylene/maleic acid (DIBMA) polymers have been extensively studied. Despite their many benefits, SMA and DIBMA polymers also have considerable drawbacks limiting their applications. Herein, we describe a series of new amphiphilic copolymers derived from DIBMA via partial amidation of the carboxylate pendant groups with various biocompatible amines. We characterize the new polymer's nanodisc-forming properties and ability to extract the melanocortin 4 receptor (MC₄R), a prototypical class A GPCR. While each new DIBMA variant displays features that may be favorable for selected applications, we identified a PEGylated DIBMA variant called mPEG₄-DIBMA as particularly promising. In the tested system mPEG₄-DIBMA abolishes unspecific interactions and outperforms other polymers by achieving higher extraction efficiencies of MC₄R from Sf9 insect cell membranes. The new nanodisc-forming polymer combines two key advantages that are crucial for investigating GPCRs in a well-defined but still native lipid-bilayer environment, thus paving the way for manifold future applications.