4-4-difluoro-4-bora-3a-4a-diaza-s-indacene and 1-palmitoyl-2-oleoylglycero-3-phosphoglycerol

Magainin 2 and PGLa are cationic, amphipathic antimicrobial peptides which when added as equimolar mixture exhibit a pronounced synergism in both their antibacterial and pore-forming activities. Here we show for the first time that the peptides assemble into defined supramolecular structures along the membrane interface. The resulting mesophases are quantitatively described by state-of-the art fluorescence self-quenching and correlation spectroscopies. Notably, the synergistic behavior of magainin 2 and PGLa correlates with the formation of hetero-domains and an order-of-magnitude increased membrane affinity of both peptides. Enhanced membrane association of the peptide mixture is only observed in the presence of phophatidylethanolamines but not of phosphatidylcholines, lipids that dominate bacterial and eukaryotic membranes, respectively. Thereby the increased membrane-affinity of the peptide mixtures not only explains their synergistic antimicrobial activity, but at the same time provides a new concept to increase the therapeutic window of combinatorial drugs.

Topics: Animals; Anti-Bacterial Agents; Antimicrobial Cationic Peptides; Boron Compounds; Cell Membrane; Drug Combinations; Drug Synergism; Ethanolamines; Fluorescent Dyes; Lipid Bilayers; Magainins; Phosphatidylcholines; Phosphatidylethanolamines; Phosphatidylglycerols; Protein Binding; Skin; Spectrometry, Fluorescence; Xenopus laevis; Xenopus Proteins

Alpha-synuclein, a presynaptic protein associated with Parkinson's disease, is found as both soluble cytosolic and membrane-bound forms. Although the function of alpha-synuclein is unknown, several observations suggest that its association with membranes is important. In the present study we investigated the effect of alpha-synuclein on lipid oxidation in membranes containing phospholipids with unsaturated fatty acids. The kinetics of lipid oxidation were monitored by the change in fluorescence intensity of the dye C11-BODIPY. We find that monomeric alpha-synuclein efficiently prevented lipid oxidation, whereas fibrillar alpha-synuclein had no such effect. Our data suggest that the prevention of unsaturated lipid oxidation by alpha-synuclein requires that it bind to the lipid membrane. The antioxidant function of alpha-synuclein is attributed to its facile oxidation via the formation of methionine sulfoxide, as shown by mass spectrometry. These findings suggest that the inhibition of lipid oxidation by alpha-synuclein may be a physiological function of the protein.

Topics: alpha-Synuclein; Antioxidants; Boron Compounds; Circular Dichroism; Fluorescence; Kinetics; Liposomes; Oxidation-Reduction; Phosphatidylcholines; Phosphatidylglycerols

Surfactant protein B (SP-B) enhances lipid insertion into the alveolar air/liquid interface upon inhalation. The aim of this study was (i) to apply a palette of tests for a detailed biochemical and biophysical characterization of SP-B and (ii) to use these tests to compare native SP-B with a fluorescent (Bodipy) SP-B analog. The method of labeling was fast and resulted in a covalent fluorophore-protein bond. The ability of both proteins to spread a surfactant film on top of a buffer surface was determined in a spreading tray using the Wilhelmy plate technique to allow detection of alterations in surface tension and calculation of spreading velocities. In a captive bubble surfactometer surface tensions of spread films were measured. Similar biophysical properties were found for both native and Bodipy-labeled SP-B. It is concluded that the combination of tests used allows detection of small differences in structure and activity between the two proteins.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Animals; Boron Compounds; Bronchoalveolar Lavage; Buffers; Cattle; Chromatography, High Pressure Liquid; Circular Dichroism; Fluorescent Dyes; Glass; Micelles; Phosphatidylcholines; Phosphatidylglycerols; Proteolipids; Pulmonary Surfactants; Pyrenes; Sequence Analysis, Protein; Spectrometry, Mass, Electrospray Ionization; Structure-Activity Relationship; Surface Tension