Browsing by Subject "ATOM FORCE-FIELD"

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  • Poojari, Chetan; Zak, Agata; Dzieciuch-Rojek, Monika; Bunker, Alex; Kepczynski, Mariusz; Rog, Tomasz (2020)
    Cholesterol plays a crucial role in modulating the physicochemical properties of biomembranes, both increasing mechanical strength and decreasing permeability. Cholesterol is also a common component of vesicle-based delivery systems, including liposome-based drug delivery systems (LDSs). However, its effect on the partitioning of drug molecules to lipid membranes is very poorly recognized. Herein, we performed a combined experimental/computational study of the potential for the use of the LDS formulation for the delivery of the antifungal drug itraconazole (ITZ). We consider the addition of cholesterol to the lipid membrane. Since ITZ is only weakly soluble in water, its bioavailability is limited. Use of an LDS has thus been proposed. We studied lipid membranes composed of cholesterol, 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphocholine (POPC), and ITZ using a combination of computational molecular dynamics (MD) simulations of lipid bilayers and Brewster angle microscopy (BAM) experiments of monolayers. Both experimental and computational results show separation of cholesterol and ITZ. Cholesterol has a strong preference to orient parallel to the bilayer normal. However, ITZ, a long and relatively rigid molecule with weakly hydrophilic groups along the backbone, predominantly locates below the interface between the hydrocarbon chain region and the polar region of the membrane, with its backbone oriented parallel to the membrane surface; the orthogonal orientation in the membrane could be the cause of the observed separation. In addition, fluorescence measurements demonstrated that the affinity of ITZ for the lipid membrane is decreased by the presence of cholesterol, which is thus probably not a suitable formulation component of an LDS designed for ITZ delivery.
  • Kulig, Waldemar; Korolainen, Hanna; Zatorska, Maria; Kwolek, Urszula; Wydro, Pawel; Kepczynski, Mariusz; Rog, Tomasz (2019)
    Phosphatidic acids (PAs) have many biological functions in biomembranes, e.g., they are involved in the proliferation, differentiation, and transformation of cells. Despite decades of research, the molecular understanding of how PAs affect the properties of biomembranes remains elusive. In this study, we explored the properties of lipid bilayers and monolayers composed of PAs and phosphatidylcholines (PCs) with various acyl chains. For this purpose, the Langmuir monolayer technique and atomistic molecular dynamics (MD) simulations were used to study the miscibility of PA and PC lipids and the molecular organization of mixed bilayers. The monolayer experiments demonstrated that the miscibility of membrane components strongly depends on the structure of the hydrocarbon chains and thus on the overall lipid shape. Interactions between PA and PC molecules vary from repulsive, for systems containing lipids with saturated and unsaturated acyl tails (strongly positive values of the excess free energy of mixing), to attractive, for systems in which all lipid tails are saturated (negative values of the excess free energy of mixing). The MD simulations provided atomistic insight into polar interactions (formation of hydrogen bonds and charge pairs) in PC-PA systems. H-bonding between PA monoanions and PCs in mixed bilayers is infrequent, and the lipid molecules interact mainly via electrostatic interactions. However, the number of charge pairs significantly decreases with the number of unsaturated lipid chains in the PA-PC system. The PA dianions weakly interact with the zwitterionic lipids, but their headgroups are more hydrated as compared to the monoanionic form. The acyl chains in all PC-PA bilayers are more ordered compared to single-component PC systems. In addition, depending on the combination of lipids, we observed a deeper location of the PA phosphate groups compared to the PC phosphate groups, which can alter the presentation of PAs for the peripheral membrane proteins, affecting their accessibility for binding.
  • Kasparyan, Gari; Poojari, Chetan; Rog, Tomasz; Hub, Jochen S. (2020)
    Itraconazole is a triazole drug widely used in the treatment of fungal infections, and it is in clinical trials for treatment of several cancers. However, the drug suffers from poor solubility, while experiments have shown that itraconazole delivery in liposome nanocarriers improves both circulation half-life and tissue distribution. The drug release mechanism from the nanocarrier is still unknown, and it depends on several factors including membrane stability against defect formation. In this work, we used molecular dynamics simulations and potential of mean force (PMF) calculations to quantify the influence of itraconazole on pore formation over lipid membranes, and we compared the effect by itraconazole with a pore-stabilizing effect by the organic solvent dimethyl sulfoxide (DMSO). According to the PMFs, both itraconazole and DMSO greatly reduce the free energy of pore formation, by up to similar to 20 kJ mol(-1). However, whereas large concentrations of itraconazole of 8 mol % (relative to lipid) were required, only small concentrations of a few mole % DMSO (relative to water) were sufficient to stabilize pores. In addition, itraconazole and DMSO facilitate pore formation by different mechanisms. Whereas itraconazole predominantly aids the formation of a partial defect with a locally thinned membrane, DMSO mainly stabilizes a transmembrane water needle by shielding it from the hydrophobic core. Notably, the two distinct mechanisms act cooperatively upon adding both itraconazole and DMSO to the membrane, as revealed by an additional reduction of the pore free energy. Overall, our simulations reveal molecular mechanisms and free energies of membrane pore formation by small molecules. We suggest that the stabilization of a locally thinned membrane as well as the shielding of a transmembrane water needle from the hydrophobic membrane core may be a general mechanism by which amphiphilic molecules facilitate pore formation over lipid membranes at sufficient concentrations.
  • Wilkosz, Natalia; Rissanen, Sami; Cyza, Malgorzata; Szybka, Renata; Nowakowska, Maria; Bunker, Alex; Rog, Tomasz; Kepczynski, Mariusz (2017)
    Uptake of piroxicam, a non-steroidal anti-inflammatory drug, from the intestines after oral intake is limited due to its low solubility and its wide use is associated with several side effects related to the gastrointestinal tract. In this study, all-atom molecular dynamics (MD) simulations and fluorescent spectroscopy were employed to investigate the interaction of piroxicam in neutral, zwitterionic, and cationic forms with lipid bilayers composed of phosphatidylcholine, cholesterol, and PEGylated lipids. Our study was aimed to assess the potential for encapsulation of piroxicam in liposomal carriers and to shed more light on the process of gastrointestinal tract injury by the drug. Through both the MD simulations and laser scanning confocal microscopy, we have demonstrated that all forms of piroxicam can associate with the lipid bilayers and locate close to the water-membrane interface. Conventional liposomes used in drug delivery are usually stabilized by the addition of cholesterol and have their bloodstream lifetime extended through the inclusion of PEGylated lipids in the formulation to create a protective polymer corona. For this reason, we tested the effect of these two modifications on the behavior of piroxicam in the membrane. When the bilayer was PEGylated, piroxicam localize to the PEG layer and within the lipid headgroup region. This suggests that PEGylated liposomes are capable of carrying a larger quantity of piroxicam than the conventional ones. (C) 2017 Elsevier B.V. All rights reserved.
  • Kepczynski, Mariusz; Rog, Tomasz (2016)
    Synthetic lipids and surfactants that do not exist in biological systems have been used for the last few decades in both basic and applied science. The most notable applications for synthetic lipids and surfactants are drug delivery, gene transfection, as reporting molecules, and as support for structural lipid biology. In this review, we describe the potential of the synergistic combination of computational and experimental methodologies to study the behavior of synthetic lipids and surfactants embedded in lipid membranes and liposomes. We focused on select cases in which molecular dynamics simulations were used to complement experimental studies aiming to understand the structure and properties of new compounds at the atomistic level. We also describe cases in which molecular dynamics simulations were used to design new synthetic lipids and surfactants, as well as emerging fields for the application of these compounds. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Rog. (C) 2016 Elsevier B.V. All rights reserved.
  • Mobarak, Edouard; Håversen, Liliana; Manna, Moutusi; Rutberg, Mikael; Levin, Malin; Perkins, Rosie; Rog, Tomasz; Vattulainen, Ilpo; Boren, Jan (2018)
    Toll-like receptor 4 (TLR4) is activated by bacterial lipopolysaccharide (LPS), which drives the production of proinflammatory cytokines. Earlier studies have indicated that cholesterol-and glycosphingolipid-rich subregions of the plasma membrane (lipid domains) are important for TLR4-mediated signaling. We report that inhibition of glucosylceramide (GluCer) synthase, which resulted in decreased concentrations of the glycosphingolipid GluCer in lipid domains, reduced the LPS-induced inflammatory response in both mouse and human macrophages. Atomistic molecular dynamics simulations of the TLR4 dimer complex (with and without LPS in its MD-2 binding pockets) in membranes (in the presence and absence of GluCer) showed that: (1) LPS induced a tilted orientation of TLR4 and increased dimer integrity; (2) GluCer did not affect the integrity of the LPS/TLR4 dimer but reduced the LPS-induced tilt; and (3) GluCer increased electrostatic interactions between the membrane and the TLR4 extracellular domain, which could potentially modulate the tilt. We also showed that GCS inhibition reduced the interaction between TLR4 and the intracellular adaptor protein Mal. We conclude that the GluCer-induced effects on LPS/TLR4 orientation may influence the signaling capabilities of the LPS/TLR4 complex by affecting its interaction with downstream signaling proteins.
  • Mastrotto, Francesca; Brazzale, Chiara; Bellato, Federica; De Martin, Sara; Grange, Guillaume; Mahmoudzadeh, Mohamad; Magarkar, Aniket; Bunker, Alex; Salmaso, Stefano; Caliceti, Paolo (2020)
    The colloidal stability, in vitro toxicity, cell association, and in vivo pharmacokinetic behavior of liposomes decorated with monomethoxy-poly(ethylene glycol)-lipids (mPEG-lipids) with different chemical features were comparatively investigated. Structural differences of the mPEG-lipids used in the study included: (a) surface-anchoring moiety [1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE), cholesterol (Chol), and cholane (Chin)]; (b) mPEG molecular weight (2 kDa mPEG and 5 kDa mPEG(114)); and (c) mPEG shape (linear and branched PEG). In vitro results demonstrated that branched (mPEG(114))(2)-DSPE confers the highest stealth properties to liposomes (similar to 31-fold lower cell association than naked liposomes) with respect to all PEGylating agents tested. However, the pharmacokinetic studies showed that the use of cholesterol as anchoring group yields PEGylated liposomes with longer permeance in the circulation and higher systemic bioavailability among the tested formulations. Liposomes decorated with mPEG(114)-Chol had 3.2- and similar to 2.1-fold higher area under curve (AUC) than naked liposomes and branched (mPEG(114))(2)-DSPE-coated liposomes, respectively, which reflects the high stability of this coating agent. By comparing the PEGylating agents with same size, namely, linear 5 kDa PEG derivatives, linear mPEG(114)-DSPE yielded coated liposomes with the best in vitro stealth performance. Nevertheless, the in vivo AUC of liposomes decorated with linear mPEG(114)-DSPE was lower than that obtained with liposomes decorated with linear mPEG(114)-Chol. Computational molecular dynamics modeling provided additional insights that complement the experimental results.
  • Koivuniemi, Artturi; Fallarero, Adyary; Bunker, Alex (2019)
    The development of antimicrobial agents that target and selectively disrupt biofilms is a pressing issue since, so far, no antibiotics have been developed that achieve this effectively. Previous experimental work has found a promising set of antibacterial peptides: β2,2-amino acid derivatives, relatively small molecules with common structural elements composed of a polar head group and two non-polar hydrocarbon arms. In order to develop insight into possible mechanisms of action of these novel antibacterial agents, we have performed an in silico investigation of four leading β2,2-amino acid derivatives, interacting with models of both bacterial (target) and eukaryotic (host) membranes, using molecular dynamics simulation with a model with all-atom resolution. We found an unexpected result that could shed light on the mechanism of action of these antimicrobial agents: the molecules assume a conformation where one of the hydrophobic arms is directed downward into the membrane core while the other is directed upwards, out of the membrane and exposed above the position of the membrane headgroups; we dubbed this conformation the “can-can pose”. Intriguingly, the can-can pose was most closely linked to the choice of headgroup. Also, the compound previously found to be most effective against biofilms displayed the strongest extent of this behavior and, additionally, this behavior was more pronounced for this compound in the bacterial than in the eukaryotic membrane. We hypothesize that adopting the can-can pose could possibly disrupt the protective peptidoglycan macronet found on the exterior of the bacterial membrane.
  • Manna, Moutusi; Javanainen, Matti; Monne, Hector Martinez-Seara; Gabius, Hans-Joachim; Rog, Tomasz; Vattulainen, Ilpo (2017)
    Extracellular and cytosolic leaflets in cellular membranes are distinctly different in lipid composition, yet they contribute together to signaling across the membranes. Here we consider a mechanism based on long-chain gangliosides for coupling the extracellular and cytosolic membrane leaflets together. Based on atomistic molecular dynamics simulations, we find that long-chain GM1 in the extracellular leaflet exhibits a strong tendency to protrude into the opposing bilayer leaflet. This interdigitation modulates the order in the cytosolic monolayer and thereby strengthens the interaction and coupling across a membrane. Coarse-grained simulations probing longer time scales in large membrane systems indicate that GM1 in the extracellular leaflet modulates the phase behavior in the cytosolic monolayer. While short-chain GM1 maintains phase-symmetric bilayers with a strong membrane registration effect, the situation is altered with long-chain GM1. Here, the significant interdigitation induced by long-chain GM1 modulates the behavior in the cytosolic GM1-free leaflet, weakening and slowing down the membrane registration process. The observed physical interaction mechanism provides a possible means to mediate or foster transmembrane communication associated with signal transduction. (C) 2017 Elsevier B.V. All rights reserved.
  • Mahmoudzadeh, Mohammad; Magarkar, Aniket; Koivuniemi, Artturi; Róg, Tomasz; Bunker, Alex (2021)
    Liposome-based drug delivery systems composed of DOPE stabilized with cholesteryl hemisuccinate (CHMS) have been proposed as a drug delivery mechanism with pH-triggered release as the anionic form (CHSa) is protonated (CHS) at reduced pH; PEGylation is known to decrease this pH sensitivity. In this manuscript, we set out to use molecular dynamics (MD) simulations with a model with all-atom resolution to provide insight into why incorporation of poly(ethyleneglycol) (PEG) into DOPE–CHMS liposomes reduces their pH sensitivity; we also address two additional questions: (1) How CHSa stabilizes DOPE bilayers into a lamellar conformation at a physiological pH of 7.4? and (2) how the change from CHSa to CHS at acidic pH triggers the destabilization of DOPE bilayers? We found that (A) CHSa stabilizes the DOPE lipid membrane by increasing the hydrophilicity of the bilayer surface, (B) when CHSa changes to CHS by pH reduction, DOPE bilayers are destabilized due to a reduction in bilayer hydrophilicity and a reduction in the area per lipid, and (C) PEG stabilizes DOPE bilayers into the lamellar phase, thus reducing the pH sensitivity of the liposomes by increasing the area per lipid through penetration into the bilayer, which is our main focus.
  • Lolicato, Fabio; Juhola, Hanna; Zak, Agata; Postila, Pekka A.; Saukko, Annina; Rissanen, Sami; Enkavi, Giray; Vattulainen, Ilpo; Kepczynski, Mariusz; Rog, Tomasz (2020)
    Synaptic neurotransmission has recently been proposed to function via either a membrane-independent or a membrane-dependent mechanism, depending on the neurotransmitter type. In the membrane-dependent mechanism, amphipathic neurotransmitters first partition to the lipid headgroup region and then diffuse along the membrane plane to their membrane-buried receptors. However, to date, this mechanism has not been demonstrated for any neurotransmitter-receptor complex. Here, we combined isothermal calorimetry measurements with a diverse set of molecular dynamics simulation methods to investigate the partitioning of an amphipathic neurotransmitter (dopamine) and the mechanism of its entry into the ligand-binding site. Our results show that the binding of dopamine to its receptor is consistent with the membrane-dependent binding and entry mechanism. Both experimental and simulation results showed that dopamine favors binding to lipid membranes especially in the headgroup region. Moreover, our simulations revealed a ligand-entry pathway from the membrane to the binding site. This pathway passes through a lateral gate between transmembrane alpha-helices 5 and 6 on the membrane-facing side of the protein. All in all, our results demonstrate that dopamine binds to its receptor by a membrane-dependent mechanism, and this is complemented by the more traditional binding mechanism directly through the aqueous phase. The results suggest that the membrane-dependent mechanism is common in other synaptic receptors, too.
  • Catte, Andrea; Girych, Mykhailo; Javanainen, Matti; Loison, Claire; Melcr, Josef; Miettinen, Markus S.; Monticelli, Luca; Maatta, Jukka; Oganesyan, Vasily S.; Ollila, O. H. Samuli; Tynkkynen, Joona; Vilov, Sergey (2016)
    Despite the vast amount of experimental and theoretical studies on the binding affinity of cations -especially the biologically relevant Na+ and Ca2+ - for phospholipid bilayers, there is no consensus in the literature. Here we show that by interpreting changes in the choline headgroup order parameters according to the 'molecular electrometer' concept [Seelig et al., Biochemistry, 1987, 26, 7535], one can directly compare the ion binding affinities between simulations and experiments. Our findings strongly support the view that in contrast to Ca2+ and other multivalent ions, Na+ and other monovalent ions (except Li+) do not specifically bind to phosphatidylcholine lipid bilayers at sub-molar concentrations. However, the Na+ binding affinity was overestimated by several molecular dynamics simulation models, resulting in artificially positively charged bilayers and exaggerated structural effects in the lipid headgroups. While qualitatively correct headgroup order parameter response was observed with Ca2+ binding in all the tested models, no model had sufficient quantitative accuracy to interpret the Ca2+: lipid stoichiometry or the induced atomistic resolution structural changes. All scientific contributions to this open collaboration work were made publicly, using nmrlipids. as the main communication platform.
  • Olzynska, Agnieszka; Kulig, Waldemar; Mikkolainen, Heikki; Czerniak, Tomasz; Jurkiewicz, Piotr; Cwiklik, Lukasz; Rog, Tomasz; Hof, Martin; Jungwirth, Pavel; Vattulainen, Ilpo (2020)
    Cholesterol renders mammalian cell membranes more compact by reducing the amount of voids in the membrane structure. Because of this, cholesterol is known to regulate the ability of cell membranes to prevent the permeation of water and water-soluble molecules through the membranes. Meanwhile, it is also known that even seemingly tiny modifications in the chemical structure of cholesterol can lead to notable changes in membrane properties. The question is, how significantly do these small changes in cholesterol structure affect the permeability barrier function of cell membranes? In this work, we applied fluorescence methods as well as atomistic molecular dynamics simulations to characterize changes in lipid membrane permeability induced by cholesterol oxidation. The studied 7 beta-hydroxycholesterol (7 beta-OH-chol) and 27-hydroxycholesterol (27-OH-chol) represent two distinct groups of oxysterols, namely, ring- and tail-oxidized cholesterols, respectively. Our previous research showed that the oxidation of the cholesterol tail has only a marginal effect on the structure of a lipid bilayer; however, oxidation was found to disturb membrane dynamics by introducing a mechanism that allows sterol molecules to move rapidly back and forth across the membranebobbing. Herein, we show that bobbing of 27-OH-chol accelerates fluorescence quenching of NBD-lipid probes in the inner leaflet of liposomes by dithionite added to the liposomal suspension. Systematic experiments using fluorescence quenching spectroscopy and microscopy led to the conclusion that the presence of 27-OH-chol increases membrane permeability to the dithionite anion. Atomistic molecular dynamics simulations demonstrated that 27-OH-chol also facilitates water transport across the membrane. The results support the view that oxysterol bobbing gives rise to successive perturbations to the hydrophobic core of the membrane, and these perturbations promote the permeation of water and small water-soluble molecules through a lipid bilayer. The observed impairment of permeability can have important consequences for eukaryotic organisms. The effects described for 27-OH-chol were not observed for 7 beta-OH-chol which represents ring-oxidized sterols.
  • Lajunen, Tatu; Nurmi, Riikka; Wilbie, Danny; Ruoslahti, Teemu; Johansson, Niklas G.; Korhonen, Ossi; Rog, Tomasz; Bunker, Alex; Ruponen, Marika; Urtti, Arto (2018)
    Light triggered drug delivery systems offer attractive possibilities for sophisticated therapy, providing both temporal and spatial control of drug release. We have developed light triggered liposomes with clinically approved indocyanine green (ICG) as the light sensitizing compound. Amphiphilic ICG can be localized in different compartments of the liposomes, but the effect of its presence, on both triggered release and long term stability, has not been studied. In this work, we report that ICG localization has a significant effect on the properties of the liposomes. Polyethylene glycol (PEG) coating of the liposomes leads to binding and stabilization of the ICG molecules on the surface of the lipid bilayer. This formulation showed both good storage stability in buffer solution (at +4-37 degrees C) and adequate stability in serum and vitreous (at +37 degrees C). The combination of ICG within the lipid bilayer and PEG coating lead to poor stability at elevated temperatures of +22 degrees C and +37 degrees C. The mechanisms of the increased instability due to ICG insertion in the lipid bilayer was elucidated with molecular dynamics simulations. Significant PEG insertion into the bilayer was induced in the presence of ICG in the lipid bilayer. Finally, feasibility of freeze-drying as a long term storage method for the ICG liposomes was demonstrated. Overall, this is the first detailed study on the interactions of lipid bilayer, light sensitizer (ICG) and PEG coating on the liposome stability. The localization of the light triggering agent significantly alters the structure of the liposomes and it is important to consider these aspects in triggered drug delivery system design.