De novo designed self-assembling helicomimetic lipooligoureas with antibacterial activity.

The overuse of antibiotics has led to a rise in infections caused by multidrug-resistant bacteria, resulting in a need for new antibacterial compounds with different modes of action. In this paper, we describe a new class of compounds called lipooligoureas, which are foldamer-based mimetics of antimicrobial lipopeptides. The lipooligoureas consist of an acyl chain connected to the N-terminus of an oligourea head group that exhibits a well-defined 2.5-helix secondary structure, which is further stabilized by the attachment of the lipophilic chain to the oligourea moiety. These compounds meet the established criteria for membranolytic compounds by possessing an amphiphilic structure that promotes the internalization and partitioning of the molecules into the lipid membrane. The presence of positively charged urea residues promotes electrostatic interactions with the negatively charged bacterial membrane. The subtle structural differences in oligourea head group influence the compounds' aggregation behavior, with the number and position of positively charged urea residues correlating with their aggregation ability. The biological activity of these compounds in inhibiting bacterial growth is correlated with their ability to aggregate, with stronger antibacterial properties exhibited by those that aggregate more easily. However, the concentration inhibiting bacterial growth is significantly lower than the critical aggregation concentration values, suggesting that the mechanism of action involves the monomeric forms of lipooligoureas. Nonetheless, a mechanism based on membrane-induced aggregation cannot be ruled out. The lipooligoureas exhibit higher activity towards Gram-positive bacteria than against Gram-negative bacteria, which is indicative of certain selectivity of these compounds. It is also demonstrated that lipooligoureas exhibit increased stability against proteolytic degradation in human blood serum.

Physicochemical and Biological Characterization of Novel Membrane-Active Cationic Lipopeptides with Antimicrobial Properties.

We have designed and synthesized new short lipopeptides composed of tetrapeptide conjugated to fatty acids with different chain lengths. The amino acid sequence of the peptide moiety included d-phenylalanine, two residues of l-2,4-diaminobutyric acid and l-leucine. To explore the possible mechanism of lipopeptide action, we have provided a physicochemical characterization of their interactions with artificial lipid membranes. For this purpose, we have used monolayers and bilayers composed of lipids representative of Gram-negative and Gram-positive bacterial membranes. Using surface pressure measurements and atomic force microscopy, we were able to monitor the changes occurring within the films upon exposure to lipopeptides. Our experiments revealed that all lipopeptides can penetrate the lipid membranes and affect their molecular ordering. The latter results in membrane thinning and fluidization. However, the effect is stronger in the lipid films mimicking Gram-positive bacterial membranes. The results of the physicochemical characterization were compared with the biological activity of lipopeptides. The effect of lipopeptides on bacterial growth was tested on several strains of bacteria. It was revealed that lipopeptides show stronger antimicrobial activity against Gram-positive bacteria. At the same time, all tested compounds display relatively low hemolytic activity.

Physicochemical Characterization of Daptomycin Interaction with Negatively Charged Lipid Membranes.

Daptomycin is known as an effective antibiotic lipopeptide which shows activity against the number of Gram-positive pathogens. Its primary target is the bacterial cell membrane. However, the detailed mechanism of daptomycin action is still subject to debate. In this paper, we have investigated the interactions between lipopeptide and model lipid films composed of negatively charged phosphatidylglycerols and cardiolipin. In order to evaluate the effect of daptomycin on the molecular organization and the properties of lipid assemblies, we have used surface pressure measurements and electrochemical methods combined with atomic force microscopy, quartz crystal microbalance, and surface-enhanced infrared absorption spectroscopy. Our results indicate that daptomycin interaction with the lipid membrane is complex. It involves daptomycin aggregation and partial insertion, which in turn affect the charge distribution on both sides of the membrane and may result in a gradient of water chemical potential. The latter can drive the flux of water across the membrane.

The shape of lipid molecules affects potential-driven molecular-scale rearrangements in model cell membranes on electrodes.

Planar asymmetric lipid bilayers composed of phosphatidylethanolamine and phosphatidylglycerol lipids are transferred onto a gold electrode surface. Lipids containing two saturated, one monounsaturated and two monounsaturated hydrocarbon chains compose the model membranes. Results of electrochemically controlled polarization modulation infrared reflection absorption spectroscopy and quartz crystal microbalance with energy dissipation studies reveal two different types of electric potential-dependent structural rearrangements in the bilayers. They are correlated with the geometry of the lipid molecule. Packing parameter correlates the cross-section area of the hydrophobic and hydrophilic parts of amphiphilic molecules. In bilayers composed of lipids with the packing parameter <1, the hydrocarbon chains are tilted with respect to the bilayer plane and the polar head groups are well hydrated. At a threshold potential an abrupt flow of water through the bilayer is connected with membrane dehydration and upward orientation of the chains. In bilayers composed of lipids with packing parameter ≥1, electric potentials have negligible effect on the membrane structure. A simple rule correlating the packing parameter with molecular scale changes occurring at electrified membranes has a large diagnostic implication for biomimetic studies and our understanding of molecular processes occurring in biological cell membranes.

Water Structure in the Submembrane Region of a Floating Lipid Bilayer: The Effect of an Ion Channel Formation and the Channel Blocker.

The structure of water in the submembrane region of the bilayer of DPhPC floating (fBLM) on a monolayer of 1-thio-ß-d-glucose (ß-Tg)-modified gold nanoparticle film was studied by the surface-enhanced infrared absorption spectroscopy (SEIRAS). SEIRAS employs surface enhancement of the mean square electric field of the photon, which is acting on a few molecular layers above the film of gold nanoparticles. Therefore, it is uniquely suited to probe water molecules in the submembrane region and provides unique information concerning the structure of the hydrogen bond network of water surrounding the lipid bilayer. The IR spectra indicated that water with a strong hydrogen network is separating the membrane from the gold surface. This water is more ordered than the water in the bulk. When alamethicin, a peptide forming ion channels, is inserted into the membrane, the network is only slightly loosened. The addition of amiloride, an ion channel blocker, results in a significant decrease in the amount of water in the submembrane region. The remaining water has a significantly distorted hydrogen bond network. This study provides unique information about the effect of the ion channel on water transport across the bilayer. The electrode potential has a relatively small effect on water structure in the submembrane region. However, the IR studies demonstrated that water is less ordered at positive transmembrane potentials. The present results provide significant insight into the nature of hydration of a floating lipid bilayer on the gold electrode surface.

Effect of Interfacial Water on the Nanomechanical Properties of Negatively Charged Floating Bilayers Supported on Gold Electrodes.

Floating lipid bilayers composed of phosphatidylglycerols and cardiolipin were deposited on gold electrodes premodified with 1-thio-ß-d-glucose monolayer by spreading of small unilamellar vesicles. The resulting lipid membrane was homogeneous, and its thickness was ∼5.0 nm. Electrochemical characterization combined with surface-enhanced infrared absorption spectroscopy revealed that negative polarization of the electrode leads to accumulation of water molecules in the interfacial region between lipid membrane and the thioglucose film. Moreover, the buildup of water layer was demonstrated to affect the nanomechanical properties of the membrane. The latter was manifested by well-pronounced decrease of Young's modulus of the lipid bilayer correlating with increasing hydration. This effect was ascribed to the decoupling of the membrane from supporting thioglucose film due to the accumulation of interfacial water. As a result, the effective stiffness of the supporting layer is lower and it alters the nanomechanical behavior of lipid membrane. Our results provide strong experimental proof for the correlation between elastic properties of floating lipid membrane and the amount of water accumulated in the submembrane region.

An in situ spectroelectrochemical study on the orientation changes of an [FeiiiLN2O3] metallosurfactant deposited as LB Films on gold electrode surfaces.

In this paper we analyze the changes in molecular orientation triggered by electrochemical reduction of an iron-containing surfactant in Langmuir-Blodgett films deposited onto gold electrodes. The metallosurfactant [Feiii(LN2O3)] (1) is an established molecular rectifier capable of unidirectional electron transfer between two electrodes. A gradual decrease in the activity is observed in sequential current vs. potential curves upon repeated cycles. Here we evaluate the redox response associated with the reduction of the Feiii/Feii couple in a single monolayer, as well as in a 5-layer LB film of 1. We use polarization modulation infrared reflection absorption spectroscopy (PM IRRAS) to follow structural and orientation changes associated with such applied potential scans. We observe that the reduction of the Fe center becomes increasingly irreversible because an Fe-Ophenolate bond is cleaved. This transformation is accompanied by an almost vertical change in the orientation of metallosurfactant molecules in LB films.