title: Structure and function of papiliocin with antimicrobial and anti-inflammatory activities isolated from the swallowtail butterfly, Papilio xuthus.
abstract: Papiliocin is a novel 37-residue cecropin-like peptide isolated recently from the swallowtail butterfly, Papilio xuthus. With the aim of identifying a potent antimicrobial peptide, we tested papiliocin in a variety of biological and biophysical assays, demonstrating that the peptide possesses very low cytotoxicity against mammalian cells and high bacterial cell selectivity, particularly against Gram-negative bacteria as well as high anti-inflammatory activity. Using LPS-stimulated macrophage RAW264.7 cells, we found that papiliocin exerted its anti-inflammatory activities by inhibiting nitric oxide (NO) production and secretion of tumor necrosis factor (TNF)-α and macrophage inflammatory protein (MIP)-2, producing effects comparable with those of the antimicrobial peptide LL-37. We also showed that the innate defense response mechanisms engaged by papiliocin involve Toll-like receptor pathways that culminate in the nuclear translocation of NF-κB. Fluorescent dye leakage experiments showed that papiliocin targets the bacterial cell membrane. To understand structure-activity relationships, we determined the three-dimensional structure of papiliocin in 300 mm dodecylphosphocholine micelles by NMR spectroscopy, showing that papiliocin has an α-helical structure from Lys(3) to Lys(21) and from Ala(25) to Val(36), linked by a hinge region. Interactions between the papiliocin and LPS studied using tryptophan blue-shift data, and saturation transfer difference-NMR experiments revealed that Trp(2) and Phe(5) at the N-terminal helix play an important role in attracting papiliocin to the cell membrane of Gram-negative bacteria. In conclusion, we have demonstrated that papiliocin is a potent peptide antibiotic with both anti-inflammatory and antibacterial activities, and we have laid the groundwork for future studies of its mechanism of action.
authors.affiliation: Department of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 143-701.
authors.forename: Jin-Kyoung
authors.initials: JK
authors.lastname: Kim
authors.affiliation: Department of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 143-701.
authors.forename: Eunjung
authors.initials: E
authors.lastname: Lee
authors.affiliation: Department of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 143-701.
authors.forename: Soyoung
authors.initials: S
authors.lastname: Shin
authors.affiliation: Department of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 143-701.
authors.forename: Ki-Woong
authors.initials: KW
authors.lastname: Jeong
authors.affiliation: Department of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 143-701.
authors.forename: Jee-Young
authors.initials: JY
authors.lastname: Lee
authors.affiliation: Laboratory of Cytokine Immunology, Institute of Biomedical Science and Technology, College of Medicine, Konkuk University, Seoul 143-701.
authors.forename: Su-Young
authors.initials: SY
authors.lastname: Bae
authors.affiliation: Laboratory of Cytokine Immunology, Institute of Biomedical Science and Technology, College of Medicine, Konkuk University, Seoul 143-701.
authors.forename: Soo-Hyun
authors.initials: SH
authors.lastname: Kim
authors.affiliation: School of Life Sciences and Biotechnology, College of Natural Sciences, Kyungpook National University, Daegu 702-701.
authors.forename: Juneyoung
authors.initials: J
authors.lastname: Lee
authors.affiliation: National Academy of Agricultural Science, Rural Development Administration, Suwon 441-100, South Korea.
authors.forename: Seong Ryul
authors.initials: SR
authors.lastname: Kim
authors.affiliation: School of Life Sciences and Biotechnology, College of Natural Sciences, Kyungpook National University, Daegu 702-701.
authors.forename: Dong Gun
authors.initials: DG
authors.lastname: Lee
authors.affiliation: National Academy of Agricultural Science, Rural Development Administration, Suwon 441-100, South Korea.
authors.forename: Jae-Sam
authors.initials: JS
authors.lastname: Hwang
authors.affiliation: Department of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 143-701. Electronic address: ymkim@konkuk.ac.kr.
authors.forename: Yangmee
authors.initials: Y
authors.lastname: Kim
mesh_terms: D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000737:chemistry* / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry* / Q000494:pharmacology*; D001419:Bacteria / Q000254:growth & development; D002080:Butterflies / Q000737:chemistry*; D002460:Cell Line; D054426:Chemokine CXCL2 / Q000096:biosynthesis; D006801:Humans; D019476:Insect Proteins / Q000737:chemistry* / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship; D051193:Toll-Like Receptors / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D023181:Antimicrobial Cationic Peptides; D054426:Chemokine CXCL2; C404915:Cxcl2 protein, mouse; D019476:Insect Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; C551727:papiliocin protein, Papilio xuthus; D009569:Nitric Oxide