Effect of Cholesterol on the Structure and Composition of Glyco-DIBMA Lipid Particles

Abstract

Different amphiphilic co-polymers have been introduced to produce polymer–lipid particles with nanodisc structure composed of an inner lipid bilayer and polymer chains self-assembled as an outer belt. These particles can be used to stabilize membrane proteins in solution and enable their characterization by means of biophysical methods, including small-angle X-ray scattering (SAXS). Some of these co-polymers have also been used to directly extract membrane proteins together with their associated lipids from native membranes. Styrene/maleic acid and diisobutylene/maleic acid are among the most commonly used co-polymers for producing polymer–lipid particles, named SMALPs and DIBMALPs, respectively. Recently, a new co-polymer, named Glyco-DIBMA, was produced by partial amidation of DIBMA with the amino sugar N-methyl-d-glucosamine. Polymer–lipid particles produced with Glyco-DIBMA, named Glyco-DIBMALPs, exhibit improved structural properties and stability compared to those of SMALPs and DIBMALPs while retaining the capability of directly extracting membrane proteins from native membranes. Here, we characterize the structure and lipid composition of Glyco-DIBMALPs produced with either 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) or 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC). Glyco-DIBMALPs were also prepared with mixtures of either POPC or DMPC and cholesterol at different mole fractions. We estimated the lipid content in the Glyco-DIBMALPs and determined the particle structure and morphology by SAXS. We show that the Glyco-DIBMALPs are nanodisc-like particles whose size and shape depend on the polymer/lipid ratio. This is relevant for designing nanodisc particles with a tunable diameter according to the size of the membrane protein to be incorporated. We also report that the addition of >20 mol % cholesterol strongly perturbed the formation of Glyco-DIBMALPs. Altogether, we describe a detailed characterization of the Glyco-DIBMALPs, which provides relevant inputs for future application of these particles in the biophysical investigation of membrane proteins.

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

“…We have previously used the same strategy to produce Glyco-DIBMA, which offers several advantages over unmodified DIBMA, including a higher protein-extraction efficiency. 32,33 Here, we sought to explore a larger chemical space through amidation using a set of structurally diverse amines, including: α-amino acids such as L-arginine (Arg) and L-2,4-diaminobutyric acid (Dab); the hexosamine meglumine (Meg); and the oligo (ethylene oxide) tetraethyleneglycol monomethyl ether amine (mPEG 4 ). These compounds have in common that they contain at least one reactive amino group, are highly water-soluble, biocompatible, and net neutral once coupled to the polymer backbone through an amide linkage.…”

Synthesis of well‐defined methacrylate copolymers and their use for stabilizing membrane proteins

“…14,15 Moreover, DIBMA proved its efficiency on various membrane proteins and expression systems. 16 Lately, derivatives of DIBMA with sulfobetaine 17 or N-methyl-D-glucosamine 18,19 pendant groups have emerged as promising polymers.…”