Saponins Form Nonionic Lipid Nanodiscs for Protein Structural Studies by Nuclear Magnetic Resonance Spectroscopy

Abstract

Structural studies of membrane proteins in native-like environments require the development of diverse membrane mimetics. Currently there is a need for nanodiscs formed with nonionic belt molecules to avoid nonphysiological electrostatic interactions between the membrane system and protein of interest. Here, we describe the formation of lipid nanodiscs from the phospholipid DMPC and a class of nonionic glycoside natural products called saponins. The morphology, surface characteristics, and magnetic alignment properties of the saponin nanodiscs were characterized by light scattering and solid-state NMR experiments. We determined that preparing nanodiscs with high saponin/lipid ratios reduced their size, diminished their ability to spontaneously align in a magnetic field, and favored insertion of individual saponin molecules in the lipid bilayer surface. Further, purification of saponin nanodiscs allowed flipping of the orientation of aligned nanodiscs by 90°. Finally, we found that aligned saponin nanodiscs provide a sufficient alignment medium to allow the measurement of residual dipolar couplings (RDCs) in aqueous cytochrome c.

Tailoring Butyl Methacrylate/Methacrylic Acid Copolymers for the Solubilization of Membrane Proteins: The Influence of Composition and Molecular Weight

“…[41] Finally, new amphiphilic small molecules (nonpolymeric but of higher MW) for membrane protein solubilization have been introduced recently, further blurring the boundary between the originally used detergents and the newly developed (mostly low-MW) polymers. [42,43] Despite the considerable progress made, the new copolymers still share some of the drawbacks of SMA mentioned above, e.g., they incorporate the potentially problematic styrenic or maleic acid moieties or show broad MW distribution. The importance of the latter parameter is being increasingly recognized because the MW (or chain length) of the copolymer is known to impact on the material's membrane solubilization efficiency and stability of the formed nanodiscs.…”

“…[ 41 ] Finally, new amphiphilic small molecules (nonpolymeric but of higher MW) for membrane protein solubilization have been introduced recently, further blurring the boundary between the originally used detergents and the newly developed (mostly low‐MW) polymers. [ 42,43 ]…”

Detergent-Free Isolation of Membrane Proteins and Strategies to Study Them in a Near-Native Membrane Environment

“…The purified membrane proteins are reconstituted in a desired membrane mimetic for biophysical and biochemical characterization. The most commonly used membrane mimetics are detergent micelles, lipid bicelles, in meso methods, amphipols, nanodiscs, and peptidiscs [ 14 , 17 , 18 , 19 ]. Each of these mimetics requires different preparation techniques and vary in their ability to function as membrane mimetics.…”

An Acrylonitrile‐Based Copolymer Gel as an NMR Alignment Medium for Extraction of Residual Dipolar Couplings of Small Molecules in Aqueous Solution

“…Many of the early reports of application of Residual Dipolar Couplings (RDCs) to the structural analysis of organic molecules were performed in aqueous solution due to the availability of many lyotropic liquid crystal (LLC) weakly‐aligning media known to work for biopolymers [1–4] . Other water‐compatible LLC systems, more recently studied or developed by our and other groups, involve chromonic systems, [5–7] as well as graphene‐oxide, [8] 1,3,5‐benzenetricarboxamide (BTA), [9] and nanodiscs [10–12] . Among the other big family of NMR weakly‐aligning media, the polymer gels, we can find also water‐compatible media such as the universal poly(ethylene glycol) gel introduced by Luy and coworkers, the copolymeric gels developed in Griesinger and Gil groups, [13–15] or gelatin‐based gels [16–18] .…”