NMR structure of pardaxin, a pore-forming antimicrobial peptide, in lipopolysaccharide micelles: mechanism of outer membrane permeabilization.
Lipopolysaccharide (LPS), the major constituent of the outer membrane of Gram-negative bacteria, is an important element against permeability of bactericidal agents, including antimicrobial peptides. However, structural determinants of antimicrobial peptides for LPS recognition are not clearly understood. Pardaxins (Pa1, Pa2, Pa3, and Pa4) are a group of pore-forming bactericidal peptides found in the mucous glands of sole fishes. Despite having a low net positive charge, pardaxins contain a broad spectrum of antibacterial activities. To elucidate the structural basis of LPS interactions of pardaxins, herein, we report the first three-dimensional structure of Pa4 bound to LPS micelles. The binding kinetics of Pa4 with LPS is estimated using [(15)N-Leu-19] relaxation dispersion NMR experiments. LPS/Pa4 interactions are further characterized by a number of biophysical methods, including isothermal titration calorimetry, (31)P NMR, saturation transfer difference NMR, dynamic light scattering, and IR spectroscopy. In the LPS-Pa4 complex, Pa4 adopts a unique helix-turn-helix conformation resembling a "horseshoe." Interestingly, the LPS-bound structure of Pa4 shows striking differences with the structures determined in lipid micelles or organic solvents. Saturation transfer difference NMR identifies residues of Pa4 that are intimately associated with LPS micelles. Collectively, our results provide mechanistic insights into the outer membrane permeabilization by pardaxin.
Bhunia
Anirban
A
From the School of Biological Sciences, Division of Structural and Computational Biology, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 and.
Domadia
Prerna N
PN
From the School of Biological Sciences, Division of Structural and Computational Biology, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 and.
Torres
Jaume
J
From the School of Biological Sciences, Division of Structural and Computational Biology, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 and.
Hallock
Kevin J
KJ
the Department of Chemistry and Biophysics, University of Michigan, Ann Arbor, Michigan 48109-1055.
Ramamoorthy
Ayyalusamy
A
the Department of Chemistry and Biophysics, University of Michigan, Ann Arbor, Michigan 48109-1055. Electronic address: ramamoor@umich.edu.
Bhattacharjya
Surajit
S
From the School of Biological Sciences, Division of Structural and Computational Biology, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 and. Electronic address: surajit@ntu.edu.sg.
D000818:Animals; D000890:Anti-Infective Agents / Q000737:chemistry* / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002463:Cell Membrane Permeability; D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism; D029941:Fish Proteins / Q000737:chemistry / Q000378:metabolism; D005397:Fish Venoms / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008070:Lipopolysaccharides / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008823:Micelles; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017550:Spectroscopy, Fourier Transform Infrared; D013696:Temperature; D013816:Thermodynamics
D000890:Anti-Infective Agents; D029941:Fish Proteins; D005397:Fish Venoms; D008070:Lipopolysaccharides; D008823:Micelles; C017603:pardaxin