title: Mode of Ezrin-Membrane Interaction as a Function of PIP2 Binding and Pseudophosphorylation.
abstract: Ezrin, a protein of the ezrin, radixin, moesin (ERM) family, provides a regulated linkage between the plasma membrane and the cytoskeleton. The hallmark of this linkage is the activation of ezrin by phosphatidylinositol-4,5-bisphosphate (PIP2) binding and a threonine phosphorylation at position 567. To analyze the influence of these activating factors on the organization of ezrin on lipid membranes and the proposed concomitant oligomer-monomer transition, we made use of supported lipid bilayers in conjunction with atomic force microscopy and fluorescence microscopy. Bilayers doped with either PIP2 as the natural receptor lipid of ezrin or a Ni-nitrilotriacetic acid-equipped lipid to bind the proteins via their His6-tags to the lipid membrane were used to bind two different ezrin variants: ezrin wild-type and ezrin T567D mimicking the phosphorylated state. Using a combination of reflectometric interference spectroscopy, atomic force microscopy, and Förster resonance energy transfer experiments, we show that only the ezrin T567D mutant, upon binding to PIP2-containing bilayers, undergoes a remarkable conformational change, which we attribute to an opening of the conformation resulting in monomeric protein on the lipid bilayer.
authors.affiliation: Institute of Medical Biochemistry and Center for Molecular Biology of Inflammation, University of Münster, Münster, Germany.
authors.forename: Victoria
authors.initials: V
authors.lastname: Shabardina
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany.
authors.forename: Corinna
authors.initials: C
authors.lastname: Kramer
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Gerdes
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Braunger
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany.
authors.forename: Andrea
authors.initials: A
authors.lastname: Cordes
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany.
authors.forename: Jonas
authors.initials: J
authors.lastname: Schäfer
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany.
authors.forename: Ingo
authors.initials: I
authors.lastname: Mey
authors.affiliation: Institute of Medical Biochemistry and Center for Molecular Biology of Inflammation, University of Münster, Münster, Germany.
authors.forename: David
authors.initials: D
authors.lastname: Grill
authors.affiliation: Institute of Medical Biochemistry and Center for Molecular Biology of Inflammation, University of Münster, Münster, Germany. Electronic address: gerke@uni-muenster.de.
authors.forename: Volker
authors.initials: V
authors.lastname: Gerke
authors.affiliation: Institute of Organic and Biomolecular Chemistry, University of Göttingen, Göttingen, Germany; Göttingen Center for Molecular Biosciences, Göttingen, Germany. Electronic address: claudia.steinem@chemie.uni-goettingen.de.
authors.forename: Claudia
authors.initials: C
authors.lastname: Steinem
mesh_terms: D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000235:genetics; D004926:Escherichia coli; D031541:Fluorescence Resonance Energy Transfer; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008051:Lipid Bilayers / Q000737:chemistry*; D018625:Microscopy, Atomic Force; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D009154:Mutation; D009571:Nitrilotriacetic Acid / Q000031:analogs & derivatives / Q000737:chemistry; D009942:Organometallic Compounds / Q000737:chemistry; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000737:chemistry*; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013057:Spectrum Analysis; D000099149:Ezrin
chemical_list: D003598:Cytoskeletal Proteins; D049452:Green Fluorescent Proteins; D008051:Lipid Bilayers; D009571:Nitrilotriacetic Acid; D009942:Organometallic Compounds; D019269:Phosphatidylinositol 4,5-Diphosphate; D011994:Recombinant Proteins; D000099149:Ezrin; C088321:nickel nitrilotriacetic acid