title: Structure, activity and interactions of the cysteine deleted analog of tachyplesin-1 with lipopolysaccharide micelle: Mechanistic insights into outer-membrane permeabilization and endotoxin neutralization.
abstract: Tachyplesin-1, a disulfide stabilized beta-hairpin antimicrobial peptide, can be found at the hemocytes of horse shoe crab Tachypleus tridentatus. A cysteine deleted linear analog of tachyplesin-1 or CDT (KWFRVYRGIYRRR-NH2) contains a broad spectrum of bactericidal activity with a reduced hemolytic property. The bactericidal activity of CDT stems from selective interactions with the negatively charged lipids including LPS. In this work, CDT-LPS interactions were investigated using NMR spectroscopy, optical spectroscopy and functional assays. We found that CDT neutralized LPS and disrupted permeability barrier of the outer membrane. Zeta potential and ITC studies demonstrated charge compensation and hydrophobic interactions of CDT with the LPS-outer membrane, respectively. Secondary structure of the peptide was probed by CD and FT-IR experiments indicating beta-strands and/or beta-turn conformations in the LPS micelle. An ensemble of structures, determined in LPS micelle by NMR, revealed a beta-hairpin like topology of the CDT peptide that was typified by an extended cationic surface and a relatively shorter segment of hydrophobic region. Interestingly, at the non-polar face, residue R11 was found to be in a close proximity to the indole ring of W2, suggesting a cation-n type interactions. Further, saturation transfer difference (STD) NMR studies established intimate contacts among the aromatic and cationic residues of CDT with the LPS micelle. Fluorescence and dynamic light scattering experiments demonstrated that CDT imparted structural destabilization to the aggregated states of LPS. Collectively, atomic resolution structure and interactions of CDT with the outer membrane-LPS could be exploited for developing potent broad spectrum antimicrobial and anti-sepsis agents.
authors.affiliation: School of Biological Sciences, Division of Structural and Computational Biology, Nanyang Technological University, Singapore 637551, Singapore.
authors.forename: Rathi
authors.initials: R
authors.lastname: Saravanan
authors.forename: Harini
authors.initials: H
authors.lastname: Mohanram
authors.forename: Mangesh
authors.initials: M
authors.lastname: Joshi
authors.forename: Prerna N
authors.initials: PN
authors.lastname: Domadia
authors.forename: Jaume
authors.initials: J
authors.lastname: Torres
authors.forename: Christiane
authors.initials: C
authors.lastname: Ruedl
authors.forename: Surajit
authors.initials: S
authors.lastname: Bhattacharjya
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000890:Anti-Infective Agents / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002151:Calorimetry; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000187:drug effects; D002942:Circular Dichroism; D003545:Cysteine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004553:Electric Conductivity; D004731:Endotoxins / Q000737:chemistry / Q000378:metabolism; D006737:Horseshoe Crabs / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008070:Lipopolysaccharides / Q000737:chemistry* / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017384:Sequence Deletion; D017550:Spectroscopy, Fourier Transform Infrared; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D000890:Anti-Infective Agents; D023181:Antimicrobial Cationic Peptides; D004268:DNA-Binding Proteins; D004731:Endotoxins; D008070:Lipopolysaccharides; D008823:Micelles; D010455:Peptides; D010456:Peptides, Cyclic; D014409:Tumor Necrosis Factor-alpha; C057636:tachyplesin peptide, Tachypleus tridentatus; D003545:Cysteine