title: Mechanism of interaction of optimized Limulus-derived cyclic peptides with endotoxins: thermodynamic, biophysical and microbiological analysis.
abstract: On the basis of formerly investigated peptides corresponding to the endotoxin-binding domain from LALF [Limulus anti-LPS (lipopolysaccharide) factor], a protein from Limulus polyphemus, we have designed and synthesized peptides of different lengths with the aim of obtaining potential therapeutic agents against septic shock syndrome. For an understanding of the mechanisms of action, we performed a detailed physicochemical and biophysical analysis of the interaction of rough mutant LPS with these peptides by applying FTIR (Fourier-transform infrared) spectroscopy, SAXS (small-angle X-ray scattering), calorimetric techniques [DSC (differential scanning calorimetry) and ITC (isothermal titration calorimetry)] and FFTEM (freeze-fracture transmission electron microscopy). Also, the action of the peptides on bacteria of different origin in microbial assays was investigated. Using FTIR and DSC, our results indicated a strong fluidization of the lipid A acyl chains due to peptide binding, with a decrease in the endothermic melting enthalpy change of the acyl chains down to a complete disappearance in the 1:0.5 to 1:2 [LPS]:[peptide] molar ratio range. Via ITC, it was deduced that the binding is a clearly exothermic process which becomes saturated at a 1:0.5 to 1:2 [LPS]:[peptide] molar ratio range. The results obtained with SAXS indicated a drastic change of the aggregate structures of LPS into a multilamellar stack, which was visualized in electron micrographs as hundreds of lamellar layers. This can be directly correlated with the inhibition of the LPS-induced production of tumour necrosis factor alpha in human mononuclear cells, but not with the action of the peptides on bacteria.
authors.affiliation: Forschungszentrum Borstel, Leibniz-Zentrum für Medizin und Biowissenschaften, Biophysik, Parkallee 10, 23845 Borstel, Germany.
authors.forename: Jörg
authors.initials: J
authors.lastname: Andrä
authors.forename: Jörg
authors.initials: J
authors.lastname: Howe
authors.forename: Patrick
authors.initials: P
authors.lastname: Garidel
authors.forename: Manfred
authors.initials: M
authors.lastname: Rössle
authors.forename: Walter
authors.initials: W
authors.lastname: Richter
authors.forename: José
authors.initials: J
authors.lastname: Leiva-León
authors.forename: Ignacio
authors.initials: I
authors.lastname: Moriyon
authors.forename: Rainer
authors.initials: R
authors.lastname: Bartels
authors.forename: Thomas
authors.initials: T
authors.lastname: Gutsmann
authors.forename: Klaus
authors.initials: K
authors.lastname: Brandenburg
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides; D060829:Arthropod Proteins; D055592:Biophysical Phenomena; D001703:Biophysics; D002151:Calorimetry; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D019610:Cytoprotection / Q000187:drug effects; D004731:Endotoxins / Q000494:pharmacology*; D006358:Hot Temperature; D006801:Humans; D007447:Invertebrate Hormones / Q000737:chemistry* / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008081:Liposomes; D050296:Microbial Viability / Q000187:drug effects; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D010456:Peptides, Cyclic / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D044367:Phase Transition; D010743:Phospholipids / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared; D013816:Thermodynamics; D014961:X-Ray Diffraction
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D060829:Arthropod Proteins; D004731:Endotoxins; D007447:Invertebrate Hormones; D008070:Lipopolysaccharides; D008081:Liposomes; D010456:Peptides, Cyclic; D010743:Phospholipids; C034705:antilipopolysaccharide factor (Limulus)