Structure of Diisobutylene Maleic Acid Copolymer (DIBMA) and Its Lipid Particle as a “Stealth” Membrane-Mimetic for Membrane Protein Research

Abstract

The study of membrane proteins remains challenging, especially in a native membrane environment. Recently, major progress has been made using maleic acid copolymers, such as styrene maleic acid, to purify membrane proteins and study them directly with native lipids associated with the membrane. Additional maleic acid copolymers, such as diisobutylene maleic acid (DIBMA) membrane-mimetic systems, are being developed and found to have improved spectroscopic properties and pH stability. We studied DIBMA and its lipid particles in solution to better understand its assembly, without and with the lipids, to provide an insight regarding how to use it in solution for better membrane extraction. Using small-angle neutron and X-ray scattering (SANS/SAXS), we show that DIBMA organizes into structures of different size scales at various concentrations and ionic strengths. The polymer performed reasonably well under most solvent conditions except in very low concentrations and high-salt conditions that could result in limited interaction with lipids. To explore DIBMA lipid particles as a suitable membrane-mimetic system for neutron scattering studies of membrane proteins, we measured and determined the contrast-matching point of DIBMA to be ∼12% (v/v) D2O  similar to that of most protiated lipid molecules but distinct from that of regular protiated proteins  providing a natural contrast for separating their neutron scattering signals. Using SANS contrast variation, we demonstrated that the scattering from the whole lipid particle can be annihilated. Further, we determined that a well-defined lipid nanodisc structure with DIBMA was contrast-matched. These results demonstrate that the DIBMA lipid particle is an outstanding “stealth” membrane-mimetic for membrane proteins. The results provide a structural framework for understanding the organization and assembly process of the polymer itself and the lipid molecules. Such an understanding is imperative for structural techniques such as cryo-electron microscopy, nuclear magnetic resonance, small-angle scattering, and other biophysical techniques.

Mechanisms of Formation, Structure, and Dynamics of Lipoprotein Discs Stabilized by Amphiphilic Copolymers: A Comprehensive Review

“…The overall disc-shaped morphology of mature lipodiscs regardless of the type of copolymer used for their preparation has been confirmed by a number of experimental techniques, including electron microscopy, atomic force microscopy [ 90 , 91 ], and small-angle scattering [ 67 , 92 ] techniques, and computer simulations [ 8 , 93 , 94 ] (see Figure 3 ). Both experiments and computer simulations indicate that the lipid patch is wrapped in a single layer [ 3 ] of polymer similar to nucleic acids in the histone complex or membrane-scaffolding protein in nanodiscs.…”

Solid-State NMR Study to Probe the Effects of Divalent Metal Ions (Ca2+ and Mg2+) on the Magnetic Alignment of Polymer-Based Lipid Nanodiscs

“…Recent studies have demonstrated the nanodisc formation by a variety of polymers that are cationic, − anionic, ,− zwitterionic, styrene-containing, styrene-free, ,, charge-free, and metal-chelators . Some of these polymers have been shown to be stable against pH, and some of them can be used to form nanodiscs for certain pH values. ,,, This library of amphipathic polymers has significantly expanded the applications of nanodiscs. − …”

Perspective on the Effect of Membrane Mimetics on Dynamic Properties of Integral Membrane Proteins

“…Lipodisq nanoparticles or SMALPs can maintain the structural and functional integrity of membrane proteins which is very difficult for traditional membrane systems. , Lipodisq nanoparticles can provide better quality experimental data for various biophysical techniques such as NMR and EPR for studying the structural dynamics of integral membrane proteins. Several forms of SMA copolymers are available in varying sizes and ratios of styrene to maleic acids to solubilize membrane proteins. ,, Various forms of reversible addition–fragmentation chain transfer (RAFT)-synthesized SMA polymers and di-isobutylene maleic acid (DIBMA) copolymers are also available to solubilize membrane proteins. − Figure shows an illustrative example of KCNQ1 incorporated into different membrane environments. The dynamic motion of integral membrane proteins is higher in detergent micelles, while the dynamic motion is restricted in liposomes when compared to that in micelles and this motion is further restricted in nanodiscs or lipodisq nanoparticles. ,− …”