Cholesteryl Hemisuccinate Is Not a Good Replacement for Cholesterol in Lipid Nanodiscs

Abstract

Nanodiscs are suitable tool for studies of membrane proteins (MPs) due to their ability to mimic native biological membranes and several MP structures are solved in nanodiscs. Among the various cell membrane components, cholesterol (CHL) is known to regulate protein function and their concentration can reach up to 50 mol %. However, studies comprising cholesterol is challenging due to its hydrophobic nature, hence, nanodiscs with only low cholesterol concentration has been studied. To overcome the problem, cholesterol analogs with high solubility in polar solutions are often used and one of them is cholesteryl hemisuccinate (CHS). Nevertheless, in molecular dynamics (MD) simulation, this is not an obstacle. In this study, we performed MD simulations of nanodiscs containing neutral phosphatidylcholine (POPC) lipids, negatively charged phosphatidylglycerol (POPG) lipids, CHL, or negatively charged cholesterol analog, CHS. Our simulations show that CHS increases the order of lipids in nanodiscs, the effect is however weaker than CHL, and even smaller in nanodiscs. Furthermore, CHS gathered around scaffold proteins while cholesterol was uniformly distributed in the nanodiscs. Thus, nanodiscs with CHS are heterogeneous and not equivalent to nanodiscs with CHL. Finally, we also observed the increased concentration of POPG near the scaffold proteins, driven by electrostatic interactions. The MD results are experimentally validated using electron paramagnetic resonance spectroscopy. These results show that nanodiscs are in fact, complex structures not easily comparable with planar lipid bilayers.

Lipid Nanodiscs via Ordered Copolymers

“…We suspect that CHS is not a replacement for cholesterol as also suggested by simulation studies comparing the two strolls. 51 Thus, high-resolution cryo-EM structures of membrane proteins as those of hTRPM4 may not necessarily indicate that they are stabilized by cholesterol, but merely that they need a bulky molecule to stabilize them in a minimum.…”

Confinement in Nanodiscs Anisotropically Modifies Lipid Bilayer Elastic Properties

“…Given the broad utility of lipid nanodiscs, it is not surprising that their properties have been studied extensively. Molecular dynamics (MD) simulations, − performed at either atomic or coarse-grained resolution, have been particularly useful to this end, as they have enabled important inferences regarding the structural properties of lipid nanodiscs and on how interactions between the nanodisc membrane and the surrounding MSPs stabilize these structures. For example, , studies revealed that the structural properties of pure nanodiscs (i.e., without embedded proteins) are not as uniform as they are in large “infinite”-sized lipid bilayers (represented in simulations by periodically repeating membrane patches).…”

General Protocol for Constructing Molecular Models of Nanodiscs

“…Several molecular dynamics (MD) studies have been performed on NDs and related lipid particles, analyzing their self-assembly and structural properties. ,− However, setting up simulation systems of NDs is not a trivial task. In 2015, Siuda and Tieleman published a potential protocol for constructing NDs in silico , based on the crystal structure with PDB ID 1AV1 .…”