Lipid nanoparticles (LNPs) composed of ionizable cationic lipids are currently the leading systems for siRNA delivery in liver disease, with the major limitation of low siRNA release efficacy into ...the cytoplasm. Ionizable cationic lipids are known to be of critical importance in LNP structure and stability, siRNA entrapment, and endosomal disruption. However, their distribution inside the LNPs and their exact role in cytoplasmic delivery remain unclear. A recent study Kulkarni
et al.
, On the formation and morphology of lipid nanoparticles containing ionizable cationic lipids and siRNA,
ACS Nano
, 2018, 12(5), 4787-4795 on LNP-siRNA systems containing the ionizable lipid DLin-KC2-DMA (also known as KC2 with an apparent p
K
a
of
ca.
6.7) suggested that neutral KC2 segregates from other components and forms an amorphous oil droplet in the core of LNPs. In this paper, we present evidence supporting the model proposed by Kulkarni
et al.
We studied KC2 segregation in the presence of POPC using molecular dynamics simulation, deuterium NMR, SAXS, and cryo-TEM experiments, and found that neutral KC2 has a high tendency to separate from POPC dispersions. KC2 confinement, upon raising the pH during the formulation process, could result in rearrangement of the internal structure of LNPs. As interactions between cationic KC2 and anionic endosomal lipids are thought to be a key factor in cargo release, KC2 confinement inside the LNP may be responsible for the observed low release efficacy.
The distribution of ionizable amino lipids (KC2) is critical in structure of lipid nanoparticles, siRNA entrapment and endosomal release. Neutral KC2 segregates from phospholipids (POPC) and forms an oily core in the bilayer interior.
Most therapeutic agents suffer from poor solubility, rapid clearance from the blood stream, a lack of targeting, and often poor translocation ability across cell membranes. Drug/gene delivery systems ...(DDSs) are capable of overcoming some of these barriers to enhance delivery of drugs to their right place of action, e.g. inside cancer cells. In this review, we focus on nanoparticles as DDSs. Complementary experimental and computational studies have enhanced our understanding of the mechanism of action of nanocarriers and their underlying interactions with drugs, biomembranes and other biological molecules. We review key biophysical aspects of DDSs and discuss how computer modeling can assist in rational design of DDSs with improved and optimized properties. We summarize commonly used experimental techniques for the study of DDSs. Then we review computational studies for several major categories of nanocarriers, including dendrimers and dendrons, polymer-, peptide-, nucleic acid-, lipid-, and carbon-based DDSs, and gold nanoparticles. This article is part of a Special Issue entitled: Membrane Proteins edited by J.C. Gumbart and Sergei Noskov.
Display omitted
•Nanoparticles have the potential to improve drug delivery and enable precision medicine.•Nanoparticles for drug delivery can be based on many different chemistries.•We review experimental and computational approaches to study nanoparticles for drug delivery.
Inverted/reverse hexagonal (HII) phases are of special interest in several fields of research, including nanomedicine. We used molecular dynamics (MD) simulation to study HII systems composed of ...dioleoylphosphatidylethanolamine (DOPE) and palmitoyloleoylphosphatidylethanolamine (POPE) at several hydration levels and temperatures. The effect of the hydration level on several HII structural parameters, including deuterium order parameters, was investigated. We further used MD simulations to estimate the maximum hydrations of DOPE and POPE HII lattices at several given temperatures. Finally, the effect of acyl chain unsaturation degree on the HII structure was studied via comparing the DOPE with POPE HII systems. In addition to MD simulations, we used deuterium nuclear magnetic resonance (2H NMR) and small-angle X-ray scattering (SAXS) experiments to measure the DOPE acyl chain order parameters, lattice plane distances, and the water core radius in HII phase DOPE samples at several temperatures in the presence of excess water. Structural parameters calculated from MD simulations are in excellent agreement with the experimental data. Dehydration decreases the radius of the water core. An increase in hydration level slightly increased the deuterium order parameter of lipids acyl chains, whereas an increase in temperature decreased it. Lipid cylinders undulated along the cylinder axis as a function of hydration level. The maximum hydration levels of PE HII phases at different temperatures were successfully predicted by MD simulations based on a single experimental measurement for the lattice plane distance in the presence of excess water. An increase in temperature decreases the maximum hydration and consequently the radius of the water core and lattice plane distances. Finally, DOPE formed HII structures with a higher curvature compared to POPE, as expected. We propose a general protocol for constructing computational HII systems that correspond to the experimental systems. This protocol could be used to study HII systems composed of molecules other than the PE systems used here and to improve and validate force field parameters by using the target data in the HII phase.
We report on atomistic simulations of DPPC lipid monolayers using the CHARMM36 lipid force field (and also the Slipid force field as a control case), combined with a four-point OPC water model. The ...entire two-phase region where domains of the “liquid-condensed” (LC) phase coexist with domains of the “liquid-expanded” (LE) phase has been explored. The simulations are long enough that the complete phase-transition stage, with two domains coexisting in the monolayer, is reached in all cases. Also, system sizes used are larger than those in previous works. As expected, domains of the minority phase are elongated, emphasizing the importance of anisotropic van der Waals and/or electrostatic dipolar interactions in the monolayer plane. The molecular structure is quantified in terms of distribution functions for the hydrocarbon chains and the PN dipoles. In contrast to previous work, where average distributions are calculated, distributions are here extracted for each of the coexisting phases by first identifying lipid molecules that belong to either LC or LE regions. In the case of the CHARMM36 force field, the three-dimensional distributions show that the average tilt angle of the chains with respect to the normal outward direction is (39.0 ± 0.1)° in the LC phase and (48.1 ± 0.5)° in the LC phase. In the case of the PN dipoles, the distributions indicate a tilt angle of (110.8 ± 0.5)° in the LC phase and (112.5 ± 0.5)° in the LE phase. These results are quantitatively different from those in previous works, which indicated a smaller normal component of the PN dipole. Also, the distributions of the monolayer-projected chains and PN dipoles have been calculated. Chain distributions peak along a particular direction in the LC domains, while they are uniform in the LE phase. Long-range ordering associated with the projected PN dipoles is absent in both phases. These results strongly suggest that LC domains do not exhibit dipolar ordering in the plane of the monolayer, the effect of these components being averaged out at short distances. Therefore, the only relevant component of the molecular dipoles, with regard to both intra- and long-range interdomain interactions, is normal to the monolayer. Also, the local orientation of chain projections is almost constant in LC domains and points in the direction along which domains are elongated, suggesting that the line tension driving the phase transition might be anisotropic with respect to the interfacial domain boundary.
Using an atom based force field, molecular dynamics (MD) simulations of 54 dodecylphosphocholine (DPC) surfactant molecules in water at two different concentrations above the critical micelle ...concentration have been performed. Starting from a random distribution of surfactants, we observed the spontaneous aggregation of the surfactants into a single micelle. At the higher DPC concentration (0.46 M) the surfactants aggregated into a worm-like micelle within 1 ns, whereas at lower concentration (0.12 M) they aggregated on a slower time scale (∼12 ns) into a spherical micelle. The difference in the final aggregate is a direct consequence of the system achieving the lowest free energy configuration for a given quantity of surfactant within the periodic boundary conditions. The simulation at low surfactant concentration was repeated three times in order to obtain statistics on the rate of aggregation. It was found that the aggregation occurs at a (virtually) constant rate with a rate constant of k = 1 × 10-4 ps-1. This is an unexpected result. On the basis of Monte Carlo simulations of a stochastic description of the system, using diffusion rates and cluster radii as determined by separate MD simulations of single DPC clusters, a lower rate constant which diminishes in the course of the aggregation process had been predicted. Neglect of hydrodynamic interactions, of long-range hydrophobic interactions, or of spatial correlations in the stochastic approach might account for the descrepancies with the more accurate MD simulations.
In this paper we study the properties of pores formed by OmpF porin from
Escherichia coli, based on a molecular dynamics simulation of the OmpF trimer, 318 palmitoyl-oleoyl-phosphatidylethanolamine ...lipids, 27 Na
+ ions, and 12,992 water molecules. After equilibration and a nanosecond production run, the OmpF trimer exhibits a C-
α root mean square deviation from the crystal structure of 0.23
nm and a stable secondary structure. No evidence is found for large-scale motions of the L3 loop. We investigate the pore dimensions, conductance, and the properties of water inside the pore. This water forms a complicated pattern, even when averaged over 1
ns of simulation time. Around the pore constriction zone the water dipoles are highly structured in the plane of the membrane, oriented by the strong transversal electric field. In addition, there is a net orientation along the pore axis pointing from the extracellular to the intracellular side of the bilayer. The diffusion coefficients of water inside the pore are greatly reduced compared to bulk. We compare our results to results from model pores (Breed et al., 1996.
Biophys. J. 70:1643–1661; Sansom et al. 1997.
Biophys. J. 73:2404–2415) and discuss implications for further theoretical work.
Certain pathogenic bacteria produce and release toxic peptides to ensure either nutrient availability or evasion from the immune system. These peptides are also toxic to the producing bacteria that ...utilize dedicated ABC transporters to provide self‐immunity. The ABC transporter McjD exports the antibacterial peptide MccJ25 in Escherichia coli. Our previously determined McjD structure provided some mechanistic insights into antibacterial peptide efflux. In this study, we have determined its structure in a novel conformation, apo inward‐occluded and a new nucleotide‐bound state, high‐energy outward‐occluded intermediate state, with a defined ligand binding cavity. Predictive cysteine cross‐linking in E. coli membranes and PELDOR measurements along the transport cycle indicate that McjD does not undergo major conformational changes as previously proposed for multi‐drug ABC exporters. Combined with transport assays and molecular dynamics simulations, we propose a novel mechanism for toxic peptide ABC exporters that only requires the transient opening of the cavity for release of the peptide. We propose that shielding of the cavity ensures that the transporter is available to export the newly synthesized peptides, preventing toxic‐level build‐up.
Synopsis
Bacteria employ dedicated ABC transporters to secrete antibacterial peptides. X‐ray structure analysis shows that the antibacterial lasso peptide transporter McjD, in contrast to other ABC transporters, lacks a stable open cavity conformation, thus preventing the influx of the exported toxic peptide.
The structure of the ABC transporter McjD was determined in two distinct conformations, apo and nucleotide‐bound.
Both conformations show an occluded cavity at both sides of the membrane.
Pulsed electron‐electron double resonance (PELDOR) measurements in the presence of the antibacterial peptide MccJ25 did not identify a stable open‐conformation.
Cross‐linking studies in proteoliposomes show that the cavity opens transiently to release the bound substrate.
Transient opening of the antibacterial peptide transporter McjD prevents the influx of its toxic substrate MccJ25 and distinguishes it from other multi‐drug transporters.
Interactions between transmembrane helices play a key role in almost all cellular processes involving membrane proteins. We have investigated helix-helix interactions in lipid bilayers with synthetic ...tryptophan-flanked peptides that mimic the membrane spanning parts of membrane proteins. The peptides were functionalized with pyrene to allow the self-association of the helices to be monitored by pyrene fluorescence and Trp-pyrene fluorescence resonance energy transfer (FRET). Specific labeling of peptides at either their N or C terminus has shown that helix-helix association occurs almost exclusively between antiparallel helices. Furthermore, computer modeling suggested that antiparallel association arises primarily from the electrostatic interactions between alpha-helix backbone atoms. We propose that such interactions may provide a force for the preferentially antiparallel association of helices in polytopic membrane proteins. Helix-helix association was also found to depend on the lipid environment. In bilayers of dioleoylphosphatidylcholine, in which the hydrophobic length of the peptides approximately matched the bilayer thickness, association between the helices was found to require peptide/lipid ratios exceeding 1/25. Self-association of the helices was promoted by either increasing or decreasing the bilayer thickness, and by adding cholesterol. These results indicate that helix-helix association in membrane proteins can be promoted by unfavorable protein-lipid interactions.
We simulated micelles of 40 (M40), 54 (M54), and 65 (M65) dodecylphosphocholine (DPC) lipids in water for up to 15 ns and analyzed the system energetics, structure of the water/lipid interface, ...structure and dynamics of the lipid tails, and overall size and shape of the micelles. M54 and M65 are similar, being mostly spherical in shape with comparable tail order parameters, atom distributions, and solvent accessible areas, whereas M40 assumes a prolate ellipsoid shape with a larger hydrophobic solvent accessible area per lipid and more restricted lipid packing. A comparison of the lipid chain structure and dynamics with those of decane and dipalmitoylphosphatidylcholine (DPPC) shows that the trans dihedral fractions are comparable, but that the dihedral transition rate is considerably slower in the micelles than in decane or DPPC, in agreement with a previous simulation of the sodium dodecyl sulfate micelle but in contrast with a recent simulation of DPC. The relaxation behavior of the CH2 segments in the lipid chains is complex, and the overall and internal motions of the lipids cannot be separated. The full orientational autocorrelation function of the CH vectors is calculated and found to decay to zero within a few nanoseconds, which is fast compared to overall micellar rotation. From a direct calculation of the spectral densities, 13C T 1 and T 2 relaxation times of the tail carbons are calculated and found to agree well with experimental measurements for the lipid chain carbons, but less well for the headgroup.
PDF
