Πέμπτη 17 Οκτωβρίου 2019

Role of Lipid Composition in the Interaction and Activity of the Antimicrobial Compound Fengycin with Complex Membrane Models

Abstract

Fengycins are compounds produced by bacteria of the Bacillus genus with strong antifungal activity. In this work, lipids extracted from fungal and oomycetal molds were used to assess the ability of fengycin to bind and insert into complex membrane models prepared as Langmuir lipid monolayers. In addition, fengycin-induced leakage in liposomes prepared from these complex lipid extracts was also evaluated. Fengycin’s ability to bind and incorporate into these membranes seemed to be mainly related to ergosterol content. Other membrane characteristics such as phospholipid fatty acyl chain length played a more peripheral role. A high ergosterol concentration appeared to allow other membrane characteristics generally associated with fengycin binding and/or insertion, such as higher proportion of phosphatidylcholine head groups or increased fatty acyl unsaturation, to be present without adversely affecting membrane integrity. Increased membrane leakage was also generally associated with the presence of low or no ergosterol. Leakage was also correlated with the previously reported biological activity of fengycin on these molds.

Effect of Triclosan on the Functioning of Liver Mitochondria and Permeability of Erythrocyte Membranes of Marsh Frog ( Pelophylax ridibundus (Pallas, 1771))

Abstract

The paper examines the effects of the antimicrobial agent triclosan on the functioning of the liver mitochondria of marsh frog (Pelophylax ridibundus (Pallas, 1771)). It was established that triclosan inhibits DNP-stimulated respiration of mitochondria and decreases respiratory control ratio. In addition, triclosan causes the collapse of the mitochondrial membrane potential on both types of substrates. Such an action of triclosan can be mediated by both a protonophore effect and suppression of the activity of complex II and combined activity of complexes II + III (and, to a lesser degree, the combined activity of complexes I + III) of the mitochondrial respiratory chain. It is shown that high concentrations of triclosan enhance the production of hydrogen peroxide during the oxidation of substrates of the complex I by mitochondria, and decrease it in the case of succinate oxidation. It is found that triclosan is able to induce nonspecific permeability of the liver mitochondria of these amphibians, as well as the plasma membrane of erythrocytes. The possible mechanisms of triclosan effect on marsh frog liver mitochondria and red blood cells are discussed.

Graphic Abstract


Prevailing Effects of Ibutilide on Fast Delayed Rectifier K + Channel

Abstract

Effects of ibutilide, a class III antiarrhythmic drug, on delayed rectifier potassium currents (IK) in freshly isolated guinea pig ventricular myocytes were studied. Experiments were performed using the whole-cell configuration of patch-clamp technique under blockade of L-type calcium currents (Cav1). Ibutilide at concentrations ranging between 10 nM and 100 µM inhibited IKr in dose-dependent manner with a half maximal effective concentration of 2.03 ± 0.74 µM (n = 5–10). The amplitude of tail currents activated by prepulse to + 20 mV was decreased from 253 ± 52 to 130 ± 25 pA (n = 8, p < 0.01) in the presence of 1 µM ibutilide. The envelope test revealed time-dependent changes in ratio of IK-tailIK during 0.2–2 s pulse durations in the absence of drug. With ibutilide, regardless of pulse duration, a relatively constant ratio was estimated, indicative of predominant involvement of IKr component. The slow IKs persisted to greater extent even at 100 μM ibutilide revealing a distinguishable selectivity toward the IKr component.

Revealing a Dual Role of Ganglioside Lipids in the Aggregation of Membrane-Associated Islet Amyloid Polypeptide

Abstract

Amyloid formation of the human islet amyloid polypeptide (hIAPP) correlates with a loss of insulin-producing beta cells in patients with type II diabetes mellitus. In this study, we investigated the binding of hIAPP to bilayers consisting of ganglioside lipids and dioleoylphosphatidylcholine (DOPC), which is a physiologically relevant lipid species for pancreatic beta cell-associated aggregation. The membrane interactions are studied computationally using a combination of coarse-grained, umbrella sampling, and atomistic molecular dynamics simulations. Herein, we demonstrate how the hIAPP peptides accumulate in the areas with a high content of ganglioside lipids. We have characterized two distinct binding modes of hIAPP on ganglioside-rich membranes, with both binding modes formed due to electrostatic interaction between the cationic peptides and the anionic ganglioside headgroup. We observed that binding in the ganglioside headgroup region induced conformational changes of the peptide towards an aggregation prone conformation, rich in β-strands. In contrast, the binding of hIAPP near the ganglioside-enriched areas mobilizes the peptide, preventing it from conformational changes and potentially shields it from interactions with other peptides. This suggests a dual role of ganglioside lipids, affecting the aggregation of hIAPP by either accelerating or inhibiting amyloid formation depending on the membrane binding and the ganglioside concentration.

Graphic Abstract


Differential Dynamics Underlying the Gln27Glu Population Variant of the β 2 -Adrenergic Receptor

Abstract

The β2-adrenergic receptor (β2AR) is a membrane-bound G-protein-coupled receptor and an important drug target for asthma. Clinical studies report that the population variant Gln27Glu is associated with a differential response to common asthma drugs, such as albuterol, isoproterenol and terbutaline. Interestingly, the 27th amino acid is positioned on the N-terminal region that is the most flexible and consequently the least studied part of the receptor. In this study, we probe the molecular origin of the differential drug binding by performing structural modeling and simulations of the wild-type (Gln) and variant (Glu) receptors followed by ensemble docking with the ligands, albuterol, isoproterenol and terbutaline. In line with clinical studies, the ligands were observed to interact preferentially with the Glu variant. Our results indicate that the Glu residue at the 27th position perturbs the network of electrostatic interactions that connects the N-terminal region to the binding site in the wild-type receptor. As a result, the Glu variant is observed to bind better to the three ligands tested in this study. Our study provides a structural basis to explain the variable drug response associated with the 27th position polymorphism in the β2AR and is a starting step to identify genotype-specific therapeutics.

Temperature Dependence of the Structure and Dynamics of a Dye-Labeled Lipid in a Planar Phospholipid Bilayer: A Computational Study

Abstract

Fluorescent probes are widely employed to label lipids for the investigation of structural and dynamic properties of model and cell membranes through optical microscopy techniques. Although the effect of tagging a lipid with an organic dye is generally assumed to be negligible, optically modified lipids can nonetheless affect the local lipid structure and, in turn, the lipid lateral mobility. To better assess this potential issue, all-atom (MD) molecular dynamics simulations have been performed to study structural and dynamic effects in a model DOPC membrane in the presence of a standard Rhodamine B-labeled DOPE lipid (RHB) as a function of temperature, i.e., 293 K, 303 K, and 320 K. As the temperature is increased, we observe similar changes in the structural properties of both pure DOPC and RHB-DOPC lipid bilayers: an increase of the area per lipid, a reduction of the membrane thickness and a decrease of lipid order parameters. The partial density profile of the RHB headgroups and their orientation within the lipid bilayer confirm the amphiphilic nature of the RHB fluorescent moiety, which mainly partitions in the DOPC glycerol backbone region at each temperature. Moreover, at all temperatures, our results on lipid lateral diffusion support a non-neutral role of the dye with respect to the unlabeled lipid mobility, thus suggesting important implications for optical microscopy studies of lipid membranes.

A Molecular Perspective on Mitochondrial Membrane Fusion: From the Key Players to Oligomerization and Tethering of Mitofusin

Abstract

Mitochondria are dynamic organelles characterized by an ultrastructural organization which is essential in maintaining their quality control and ensuring functional efficiency. The complex mitochondrial network is the result of the two ongoing forces of fusion and fission of inner and outer membranes. Understanding the functional details of mitochondrial dynamics is physiologically relevant as perturbations of this delicate equilibrium have critical consequences and involved in several neurological disorders. Molecular actors involved in this process are large GTPases from the dynamin-related protein family. They catalyze nucleotide-dependent membrane remodeling and are widely conserved from bacteria to higher eukaryotes. Although structural characterization of different family members has contributed in understanding molecular mechanisms of mitochondrial dynamics in more detail, the complete structure of some members as well as the precise assembly of functional oligomers remains largely unknown. As increasing structural data become available, the domain modularity across the dynamin superfamily emerged as a foundation for transfering the knowledge towards less characterized members. In this review, we will first provide an overview of the main actors involved in mitochondrial dynamics. We then discuss recent example of computational methodologies for the study of mitofusin oligomers, and present how the usage of integrative modeling in conjunction with biochemical data can be an asset in progressing the still challenging field of membrane dynamics.

Interdigitation of Lipids Induced by Membrane–Active Proteins

Abstract

The membrane–active protein Nogo-66 is found to induce interdigitation in dimyristoylphosphocholine membranes. Extensive molecular dynamics simulations have been employed to probe the interactions of Nogo-66 with these model membranes. This phase change happens when the temperature is close to the main transition temperature of the membrane (Tm) and only in the presence of the protein. No similar interdigitation of the membrane lipids was observed temperatures well above Tm in the presence of the protein. In addition, in protein-free simulations, no interdigitation of the membrane lipids was found both at temperatures near or well above Tm indicating that the observed effect is caused by the interactions of Nogo-66 with the membrane. Analysis of the simulations suggest protein–membrane interactions, even if transient, alter the lifetimes of lipid head defects and can potentially alter the effective Tm and cause interdigitation. This study emphasize the importance of membrane–active proteins and their interactions with membranes leading to phase transitions which would affect other membrane-related processes such as domain formation.

Effects of Post-translational Modifications on Membrane Localization and Signaling of Prostanoid GPCR–G Protein Complexes and the Role of Hypoxia

Abstract

G protein-coupled receptors (GPCRs) play a pivotal role in the adaptive responses to cellular stresses such as hypoxia. In addition to influencing cellular gene expression profiles, hypoxic microenvironments can perturb membrane protein localization, altering GPCR effector scaffolding and altering downstream signaling. Studies using proteomics approaches have revealed significant regulation of GPCR and G proteins by their state of post-translational modification. The aim of this review is to examine the effects of post-translational modifications on membrane localization and signaling of GPCR–G protein complexes, with an emphasis on vascular prostanoid receptors, and to highlight what is known about the effect of cellular hypoxia on these mechanisms. Understanding post-translational modifications of protein targets will help to define GPCR targets in treatment of disease, and to inform research into mechanisms of hypoxic cellular responses.

Graphic Abstract


Interaction of Antimicrobial Lipopeptides with Bacterial Lipid Bilayers

Abstract

The resistance of pathogens to traditional antibiotics is currently a global issue of enormous concern. As the discovery and development of new antibiotics become increasingly challenging, synthetic antimicrobial lipopeptides (AMLPs) are now receiving renewed attention as a new class of antimicrobial agents. In contrast to traditional antibiotics, AMLPs act by physically disrupting the cell membrane (rather than targeting specific proteins), thus reducing the risk of inducing bacterial resistance. In this study, we use microsecond-timescale atomistic molecular dynamics simulations to quantify the interaction of a short AMLP (C16-KKK) with model bacterial lipid bilayers. In particular, we investigate how fundamental transmembrane properties change in relation to a range of lipopeptide concentrations. A number of structural, mechanical, and dynamical features are found to be significantly altered in a non-linear fashion. At 10 mol% concentration, lipopeptides have a condensing effect on bacterial bilayers, characterized by a decrease in the area per lipid and an increase in the bilayer order. Higher AMLP concentrations of 25 and 40 mol% destabilize the membrane by disrupting the bilayer core structure, inducing membrane thinning and water leakage. Important transmembrane properties such as the lateral pressure and dipole potential profiles are also affected. Potential implications on membrane function and associated proteins are discussed.

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