title: Structural basis for endotoxin neutralization by the eosinophil cationic protein.
abstract: Acute infection by Gram-negative pathogens can induce an exacerbated immune response that leads to lethal septic shock syndrome. Bacterial lipopolysaccharide (LPS) is a major pathogen-associated molecular pattern molecule that can initiate massive and lethal immune system stimulation. Therefore, the development of new and effective LPS-neutralizing agents is a top priority. The eosinophil cationic protein (ECP) is an antimicrobial protein secreted in response to infection, with a remarkable affinity for LPS. In the present study, we demonstrate that ECP is able to neutralize bacterial LPS and inhibit tumor necrosis factor-α production in human macrophages. We also characterized ECP neutralizing activity using progressively truncated LPS mutants, and conclude that the polysaccharide moiety and lipid A portions are required for LPS-mediated neutralization. In addition, we mapped the structural determinants required for the ECP-LPS interaction by nuclear magnetic resonance. Our results show that ECP is able to neutralize LPS and therefore opens a new route for developing novel therapeutic agents based on the ECP structural scaffolding.
authors.affiliation: Department of Biochemistry and Molecular Biology, Biosciences Faculty, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Pulido
authors.affiliation: Departamento de Química Biológica, Instituto de Química-Física Rocasolano, CSIC, Madrid, Spain.
authors.forename: Maria Flor
authors.initials: MF
authors.lastname: Garcia-Mayoral
authors.affiliation: Department of Biochemistry and Molecular Biology, Biosciences Faculty, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Moussaoui
authors.affiliation: Department of Biochemistry and Molecular Biology, Biosciences Faculty, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain.
authors.forename: Diego
authors.initials: D
authors.lastname: Velázquez
authors.affiliation: Department of Biochemistry and Molecular Biology, Biosciences Faculty, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain.
authors.forename: Marc
authors.initials: M
authors.lastname: Torrent
authors.affiliation: Departamento de Química Biológica, Instituto de Química-Física Rocasolano, CSIC, Madrid, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Bruix
authors.affiliation: Department of Biochemistry and Molecular Biology, Biosciences Faculty, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain.
authors.forename: Ester
authors.initials: E
authors.lastname: Boix
mesh_terms: D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D004731:Endotoxins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D047150:Eosinophil Cationic Protein / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008070:Lipopolysaccharides / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D000072417:Protein Domains; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D004731:Endotoxins; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha; D047150:Eosinophil Cationic Protein
keywords: antimicrobial peptides; endotoxin; eosinophil cationic protein; lipopolysaccharide; structure-function