Lipid-Protein Nanodiscs Offer New Perspectives for Structural and Functional Studies of Water-Soluble Membrane-Active Peptides

Abstract

Lipid-protein nanodiscs (LPNs) are nanoscaled fragments of a lipid bilayer stabilized in solution by the apolipoprotein or a special membrane scaffold protein (MSP). In this work, the applicability of LPN-based membrane mimetics in the investigation of water-soluble membrane-active peptides was studied. It was shown that a pore-forming antimicrobial peptide arenicin-2 from marine lugworm (charge of +6) disintegrates LPNs containing both zwitterionic phosphatidylcholine (PC) and anionic phosphatidylglycerol (PG) lipids. In contrast, the spider toxin VSTx1 (charge of +3), a modifier of Kv channel gating, effectively binds to the LPNs containing anionic lipids (POPC/DOPG, 3 : 1) and does not cause their disruption. VSTx1 has a lower affinity to LPNs containing zwitterionic lipids (POPC), and it weakly interacts with the protein component of nanodiscs, MSP (charge of –6). The neurotoxin II (NTII, charge of +4) from cobra venom, an inhibitor of the nicotinic acetylcholine receptor, shows a comparatively low affinity to LPNs containing anionic lipids (POPC/DOPG, 3 : 1 or POPC/DOPS, 4 : 1), and it does not bind to LPNs/POPC. The obtained data show that NTII interacts with the LPN/POPC/DOPS surface in several orientations, and that the exchange process among complexes with different topologies proceeds fast on the NMR timescale. Only one of the possible NTII orientations allows for the previously proposed specific interaction between the toxin and the polar head group of phosphatidylserine from the receptor environment (Lesovoy et al., Biophys. J. 2009. V. 97. № 7. P. 2089–2097). These results indicate that LPNs can be used in structural and functional studies of water-soluble membrane-active peptides (probably except pore-forming ones) and in studies of the molecular mechanisms of peptide-membrane interaction.

NMR Dynamics of Transmembrane and Intracellular Domains of p75NTR in Lipid-Protein Nanodiscs

“…This presents a nice prospect for the use of LPNs as a membrane mimetic for structural studies of multidomain membrane proteins, especially since LPNs have been shown to be a perfect medium for the refolding of membrane proteins (32). Nanodiscs were first proposed as a reference medium for detergent screening (28) or as a possible environment in which to study the topology of small peptides in the membrane (33)(34)(35)(36); however, recent advances, such as the development of new MSPs that form smaller discoid particles (up to~50 kDa) (31), isotope labeling strategies, and NMR experimental techniques have also made them applicable for structural studies of membrane proteins (37). In this work, we show that a transmembrane protein with a relatively large water-soluble domain (p75-DECD) can be incorporated into LPNs of various sizes and studied by the conventional means of solution NMR spectroscopy to obtain biologically relevant structural data.…”

Quantum dot surface engineering: Toward inert fluorophores with compact size and bright, stable emission

“…However, this assembled structure is only stable when a secondary amphiphile is added that can bind to the high curvature edges. In this case, helical lipoproteins called membrane scaffolding proteins (MSPs) have been shown to efficiently bind to the sides of lamellar sheets to generate nanodisc structures that are alone highly stable [196]. This combination of phospholipids and MSPs efficiently coat all sides of a nanoplatelet, with different adsorbates localizing to different regions depending on curvature (Fig.…”

Recent advances in nanodisc technology for membrane protein studies (2012–2017)

“…Nanodiscs with different lipid composition have been used to study membrane‐bound active peptides . In a recent work, the authors showed that arenicin‐2, a pore‐forming antimicrobial peptide, disintegrates nanodiscs containing both zwitterionic phosphatidylcholine (PC) and anionic phosphatidylglycerol (PG).…”

“…This is due to the propensity of membrane proteins to aggregate outside a membrane environment during their isolation from membranes using detergents. Therefore, in recent years, several lipid membrane mimics were developed in order to facilitate interrogation of membrane proteins by rigorous biophysical and analytical techniques. In 2002, Sligar and coworkers at the University of Illinois at Urbana‐Champaign developed a new technology termed as nanodiscs based on previous experimentation with ApoA1 and synthetic phospholipids .…”