title: Membrane permeabilization, orientation, and antimicrobial mechanism of subtilosin A.
abstract: Subtilosin A is an antimicrobial peptide produced by the soil bacterium Bacillus subtilis that possesses bactericidal activity against a diverse range of bacteria, including Listeria monocytogenes. Recent structural studies have found that subtilosin A is posttranslationally modified in a unique way, placing it in a new class of bacteriocins. In this study, in order to understand the mechanism of membrane-disruption by subtilosin A, the interaction of the peptide with model phospholipid bilayers is characterized using fluorescence, solid-state NMR and differential scanning calorimetry (DSC) experiments. Our results in this study show that subtilosin A interacts with the lipid head group region of bilayer membranes in a concentration dependent manner. Fluorescence experiments reveal the interaction of subtilosin A with small unilamellar vesicles (SUVs) composed of POPC, POPG and E. coli total lipids, and that at least one edge of the molecule is buried in membrane bilayers. At high concentrations, it induces leakage from SUVs of POPC and POPE/POPG (7:3) mixture. (15)N solid-state NMR data suggests that the cyclic peptide is partially inserted into bilayers, which is in agreement with the fluorescence data. (31)P and (2)H NMR experiments and DSC data support the hypothesis that subtilosin A adopts a partially buried orientation in lipid bilayers, by showing that it induces a conformational change in the lipid headgroup and disordering in the hydrophobic region of bilayers. These results suggest that the lipid perturbation observed in this study may be one of the consequences of subtilosin A binding to lipid bilayers, which results in membrane permeabilization at high peptide concentrations.
authors.affiliation: Department of Chemistry and Biophysics Research Division, University of Michigan, Ann Arbor, MI 48109-1055, USA.
authors.forename: Sathiah
authors.initials: S
authors.lastname: Thennarasu
authors.forename: Dong-Kuk
authors.initials: DK
authors.lastname: Lee
authors.forename: Alan
authors.initials: A
authors.lastname: Poon
authors.forename: Karen E
authors.initials: KE
authors.lastname: Kawulka
authors.forename: John C
authors.initials: JC
authors.lastname: Vederas
authors.forename: Ayyalusamy
authors.initials: A
authors.lastname: Ramamoorthy
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001430:Bacteriocins / Q000737:chemistry / Q000378:metabolism; D002152:Calorimetry, Differential Scanning; D002463:Cell Membrane Permeability; D008051:Lipid Bilayers / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D010456:Peptides, Cyclic; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
chemical_list: D001426:Bacterial Proteins; D001430:Bacteriocins; D008051:Lipid Bilayers; D008081:Liposomes; D010455:Peptides; D010456:Peptides, Cyclic; C047828:sboA protein, Bacillus subtilis