title: Alanine scan and (2)H NMR analysis of the membrane-active peptide BP100 point to a distinct carpet mechanism of action.
abstract: The short membrane-active peptide BP100 [KKLFKKILKYL-NH2] is known as an effective antimicrobial and cell penetrating agent. For a functional alanine scan each of the 11 amino acids was replaced with deuterated Ala-d3, one at a time. MIC assays showed that a substitution of Lys did not affect the antimicrobial activity, but it decreased when a hydrophobic residue was replaced. In most cases, a reduction in hydrophobicity led to a decrease in hemolysis, and some peptide analogues had an improved therapeutic index. Circular dichroism showed that BP100 folds as an amphiphilic α-helix in a bilayer. Its alignment was determined from (2)H NMR in oriented membranes of different composition. The azimuthal rotation angle was the same under all conditions, but the average helix tilt angle and the dynamical behavior of the peptide varied in a systematic manner. In POPC/POPG bilayers, with a negative spontaneous curvature, the peptide was found to lie flat on the bilayer surface, and with little wobble. In DMPC/DMPG, with a positive spontaneous curvature, BP100 at higher concentrations became tilted obliquely into the membrane, with the uncharged C-terminus inserted more deeply into the lipid bilayer, experiencing significant fluctuations in tilt angle. In DMPC/DMPG/lyso-MPC, with a pronounced positive spontaneous curvature, the helix tilted even further and became even more mobile. The 11-mer BP100 is obviously too short to form transmembrane pores. We conclude that BP100 operates via a carpet mechanism, whereby the C-terminus gets inserted into the hydrophobic core of the bilayer, which leads to membrane perturbation and induces transient permeability.
authors.affiliation: Instituto de Química Física "Rocasolano", CSIC, Serrano 119, 28006 Madrid, Spain.
authors.forename: Héctor
authors.initials: H
authors.lastname: Zamora-Carreras
authors.affiliation: Karlsruhe Institute for Technology (KIT), Institute for Biological Interfaces (IBG-2), POB 3640, 76021 Karlsruhe, Germany.
authors.forename: Erik
authors.initials: E
authors.lastname: Strandberg
authors.affiliation: Karlsruhe Institute for Technology (KIT), Institute for Biological Interfaces (IBG-2), POB 3640, 76021 Karlsruhe, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Mühlhäuser
authors.affiliation: Karlsruhe Institute for Technology (KIT), Institute for Biological Interfaces (IBG-2), POB 3640, 76021 Karlsruhe, Germany.
authors.forename: Jochen
authors.initials: J
authors.lastname: Bürck
authors.affiliation: Karlsruhe Institute for Technology (KIT), Institute for Biological Interfaces (IBG-2), POB 3640, 76021 Karlsruhe, Germany.
authors.forename: Parvesh
authors.initials: P
authors.lastname: Wadhwani
authors.affiliation: Instituto de Química Física "Rocasolano", CSIC, Serrano 119, 28006 Madrid, Spain.
authors.forename: M Ángeles
authors.initials: MÁ
authors.lastname: Jiménez
authors.affiliation: Instituto de Química Física "Rocasolano", CSIC, Serrano 119, 28006 Madrid, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Bruix
authors.affiliation: Karlsruhe Institute for Technology (KIT), Institute for Biological Interfaces (IBG-2), POB 3640, 76021 Karlsruhe, Germany; KIT, Institute of Organic Chemistry and CFN, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany. Electronic address: anne.ulrich@kit.edu.
authors.forename: Anne S
authors.initials: AS
authors.lastname: Ulrich
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects; D002942:Circular Dichroism; D003903:Deuterium; D006461:Hemolysis / Q000187:drug effects; D008051:Lipid Bilayers; D008565:Membrane Proteins / Q000737:chemistry*; D008826:Microbial Sensitivity Tests; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D009842:Oligopeptides / Q000737:chemistry* / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D008051:Lipid Bilayers; D008565:Membrane Proteins; D009842:Oligopeptides; C540525:lysyl-lysyl-leucyl-phenylalanyl-lysyl-lysyl-isoleucyl-leucyl-lysyl-tyrosyl-leucinamide; D003903:Deuterium
keywords: Alanine scan; Amphipathic α-helix; Antimicrobial peptide BP100; Dynamical NMR data analysis; Geometric analysis of labeled alanines (GALA); Lipid spontaneous curvature; Peptide orientation and dynamics