Struktur-Wirkungsbeziehungen zwischen membranaktiven Peptiden und bakteriellen Lipopolysacchariden

Biophysikalische Untersuchungen zur Struktur-Wirkungsbeziehung von membranaktiven Peptiden und Lipopolysaccharide

Calcium adsorption and displacement: characterization of lipid monolayers and their interaction with membrane-active peptides/proteins

BACKGROUND: The first target of antimicrobial peptides (AMPs) is the bacterial membrane. In the case of Gram-negative bacteria this is the outer membrane (OM), the lipid composition of which is extremely asymmetric: Whereas the inner leaflet is composed of a phospholipid mixture, the outer leaflet is made up solely from lipopolysaccharides (LPSs). LPS, therefore, represents the first target of AMPs. The binding and intercalation of polycationic AMPs is driven by the number and position of negatively charged groups of the LPS. Also, proteins other than cationic AMPs can interact with LPS, e.g. leading eventually to a neutralization of the endotoxic effects of LPS. We compared different biophysical techniques to gain insight into the properties of the electrical surface potentials of lipid monolayers and aggregates composed of LPSs and various phospholipids and their interaction with peptides and proteins. RESULTS: The net negative charge calculated from the chemical structure of the phospholipid and LPS molecules is linearly correlated with the adsorption of calcium to two-dimensional lipid monolayers composed of the respective lipids. However, the ζ-potentials determined by the electrophoretic mobility of LPS aggregates can only be interpreted by assuming a dependence of the plane of shear on the number of saccharides and charged groups. Various peptides and proteins were able to displace calcium adsorbed to monolayers. CONCLUSION: To characterize the electrical properties of negatively charged phospholipids and LPSs and their electrostatic interaction with various polycationic peptides/proteins, the adsorption of calcium to and displacement from lipid monolayers is a suitable parameter. Using the calcium displacement method, the binding of peptides to monolayers can be determined even if they do not intercalate. The interpretation of ζ-potential data is difficulty for LPS aggregates, because of the complex three-dimensional structure of the LPS molecules. However, the influence of peptides/proteins on the ζ-potential can be used to characterize the underlying interaction mechanisms

DC diaphragm discharge in water solutions of selected organic acids

Effect of four simple organic acids water solution on a DC diaphragm discharge was studied. Efficiency of the discharge was quantified by the hydrogen peroxide production determined by UV-VIS spectrometry of a H2O2 complex formed with specific titanium reagent. Automatic titration was used to study the pH behaviour after the plasma treatment. Optical emission spectroscopy overview spectra were recorded and detailed spectra of OH band and Hβ line were used to calculate the rotational temperature and comparison of the line profile (reflecting electron concentration) in the acid solutions

Lipopolysaccharide interaction is decisive for the activity of the antimicrobial peptide NK-2 against Escherichia coli and Proteus mirabilis

International audiencePhosphatidylglycerol is a widely used mimetic to study the effects of antimicrobial peptides (AMPs) on the bacterial cytoplasmic membrane. It turned out, however, that the antibacterial activities of novel NK-2-derived AMPs could not sufficiently explained by using this simple model system. Since the lipopolysaccharide (LPS) containing outer membrane is the first barrier of Gram-negative bacteria, here we investigated interactions of NK-2 and a shortened variant thereof with viable Escherichia coli WBB01 and Proteus mirabilis R45, and with model membranes composed of LPS isolated from these two strains. Differences in net charge and charge distribution of the two LPS have been made responsible for the differential sensitivity of the respective bacteria to other AMPs. As imaged by TEM and AFM, NK-2-mediated killing of these bacteria was corroborated by structural alterations of the outer and inner membranes, the release of E. coli cytoplasma, and the formation of unique fibrous structures inside P. mirabilis, suggesting distinct and novel intracellular targets. NK-2 bound to and intercalated into LPS bilayers, and eventually induced the formation of transient, heterogeneous lesions in planar lipid bilayers. However, the discriminative activity of NK-2 against the two bacterial strains was independent from membrane intercalation and lesion formation, which both were indistinguishable for the two LPS. Instead, differences in activity originated from the LPS binding step, which could be demonstrated by NK 2 attachment to intact bacteria, and to solid-supported LPS bilayers on a surface acoustic wave biosensor