title: EPR and FTIR studies reveal the importance of highly ordered sterol-enriched membrane domains for ostreolysin activity.
abstract: Ostreolysin is a cytolytic protein from the edible oyster mushroom (Pleurotus ostreatus), which recognizes specifically and binds to raft-like sterol-enriched membrane domains that exist in the liquid-ordered phase. Its binding can be abolished by micromolar concentrations of lysophospholipids and fatty acids. The membrane activity of ostreolysin, however, does not completely correlate with the ability of a certain sterol to induce the formation of a liquid-ordered phase, suggesting that the protein requires an additional structural organization of the membrane to exert its activity. The aim of this study was to further characterize the lipid membranes that facilitate ostreolysin binding by analyzing their lipid phase domain structure. Fourier-transformed infrared spectroscopy (FTIR) and electron paramagnetic resonance (EPR) were used to analyze the ordering and dynamics of membrane lipids and the membrane domain structure of a series of unilamellar liposomes prepared by systematically changing the lipid components and their ratios. Our results corroborate the earlier conclusion that the average membrane fluidity of ostreolysin-susceptible liposomes alone cannot account for the membrane activity of the protein. Combined with previous data computer-aided interpretation of EPR spectra strongly suggests that chemical properties of membrane constituents, their specific distribution, and physical characteristics of membrane nanodomains, resulting from the presence of sterol and sphingomyelin (or a highly ordered phospholipid, dipalmitoylphosphatidylcholine), are essential prerequisites for ostreolysin membrane binding and pore-formation.
authors.affiliation: Department of Biology, Biotechnical Faculty, University of Ljubljana, Vecna pot 111, 1000 Ljubljana, Slovenia.
authors.forename: Katja
authors.initials: K
authors.lastname: Rebolj
authors.forename: Biserka
authors.initials: B
authors.lastname: Bakrac
authors.forename: Maja
authors.initials: M
authors.lastname: Garvas
authors.forename: Katja
authors.initials: K
authors.lastname: Ota
authors.forename: Marjeta
authors.initials: M
authors.lastname: Sentjurc
authors.forename: Cristina
authors.initials: C
authors.lastname: Potrich
authors.forename: Manuela
authors.initials: M
authors.lastname: Coraiola
authors.forename: Rossella
authors.initials: R
authors.lastname: Tomazzolli
authors.forename: Mauro
authors.initials: M
authors.lastname: Dalla Serra
authors.forename: Peter
authors.initials: P
authors.lastname: Macek
authors.forename: Kristina
authors.initials: K
authors.lastname: Sepcić
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D005656:Fungal Proteins / Q000737:chemistry; D006460:Hemolysin Proteins / Q000737:chemistry*; D008081:Liposomes / Q000737:chemistry; D021962:Membrane Microdomains / Q000737:chemistry / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared; D013109:Sphingomyelins / Q000737:chemistry; D013261:Sterols / Q000737:chemistry*; D013552:Swine
chemical_list: D005656:Fungal Proteins; D006460:Hemolysin Proteins; D008081:Liposomes; D013109:Sphingomyelins; D013261:Sterols; C461280:ostreolysin