title: Orientation of a beta-hairpin antimicrobial peptide in lipid bilayers from two-dimensional dipolar chemical-shift correlation NMR.
abstract: The orientation of a beta-sheet membrane peptide in lipid bilayers is determined, for the first time, using two-dimensional (2D) (15)N solid-state NMR. Retrocyclin-2 is a disulfide-stabilized cyclic beta-hairpin peptide with antibacterial and antiviral activities. We used 2D separated local field spectroscopy correlating (15)N-(1)H dipolar coupling with (15)N chemical shift to determine the orientation of multiply (15)N-labeled retrocyclin-2 in uniaxially aligned phosphocholine bilayers. Calculated 2D spectra exhibit characteristic resonance patterns that are sensitive to both the tilt of the beta-strand axis and the rotation of the beta-sheet plane from the bilayer normal and that yield resonance assignment without the need for singly labeled samples. Retrocyclin-2 adopts a transmembrane orientation in dilauroylphosphatidylcholine bilayers, with the strand axis tilted at 20 degrees +/- 10 degrees from the bilayer normal, but changes to a more in-plane orientation in thicker 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidyl-choline (POPC) bilayers with a tilt angle of 65 degrees +/- 15 degrees . These indicate that hydrophobic mismatch regulates the peptide orientation. The 2D spectra are sensitive not only to the peptide orientation but also to its backbone (phi, psi) angles. Neither a bent hairpin conformation, which is populated in solution, nor an ideal beta-hairpin with uniform (phi, psi) angles and coplanar strands, agrees with the experimental spectrum. Thus, membrane binding orders the retrocyclin conformation by reducing the beta-sheet curvature but does not make it ideal. (31)P NMR spectra of lipid bilayers with different compositions indicate that retrocyclin-2 selectively disrupts the orientational order of anionic membranes while leaving zwitteronic membranes intact. These structural results provide insights into the mechanism of action of this beta-hairpin antimicrobial peptide.
authors.affiliation: Department of Chemistry, Iowa State University, Ames, Iowa 50011, USA.
authors.forename: Ming
authors.initials: M
authors.lastname: Tang
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Waring
authors.forename: Robert I
authors.initials: RI
authors.lastname: Lehrer
authors.forename: Mei
authors.initials: M
authors.lastname: Hong
mesh_terms: D023181:Antimicrobial Cationic Peptides / Q000737:chemistry*; D003198:Computer Simulation; D023082:Defensins / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008560:Membrane Fluidity; D008565:Membrane Proteins / Q000737:chemistry*; D008956:Models, Chemical; D008958:Models, Molecular; D010456:Peptides, Cyclic / Q000737:chemistry*; D011487:Protein Conformation
chemical_list: D023181:Antimicrobial Cationic Peptides; D023082:Defensins; D008051:Lipid Bilayers; D008565:Membrane Proteins; D010456:Peptides, Cyclic; C511481:retrocyclin-2, human