title: Structure and dynamics of Type III periplasmic proteins VcFhuD and VcHutB reveal molecular basis of their distinctive ligand binding properties.
abstract: Molecular mechanisms of xenosiderophore and heme acquisitions using periplasmic binding protein (PBP) dependent ATP-binding cassette transporters to scavenge the essential nutrient iron are elusive yet in Vibrio cholerae. Our current study delineates the structures, dynamics and ligand binding properties of two Type III PBPs of V. cholerae, VcFhuD and VcHutB. Through crystal structures and fluorescence quenching studies we demonstrate unique features of VcFhuD to bind both hydroxamate and catecholate type xenosiderophores. Like E. coli FhuD, VcFhuD binds ferrichrome and ferri-desferal using conserved Tryptophans and R102. However, unlike EcFhuD, slightly basic ligand binding pocket of VcFhuD could favour ferri-enterobactin binding with plausible participation of R203, along with R102, like it happens in catecholate binding PBPs. Structural studies coupled with spectrophotometric and native PAGE analysis indicated parallel binding of two heme molecules to VcHutB in a pH dependent manner, while mutational analysis established the relative importance of Y65 and H164 in heme binding. MD simulation studies exhibited an unforeseen inter-lobe swinging motion in Type III PBPs, magnitude of which is inversely related to the packing of the linker helix with its neighboring helices. Small inter-lobe movement in VcFhuD or dramatic twisting in VcHutB is found to influence ligand binding.
authors.affiliation: Department of Biotechnology, St. Xavier's College, 30 Park Street, Kolkata 700016, India.
authors.forename: Shubhangi
authors.initials: S
authors.lastname: Agarwal
authors.affiliation: Department of Biotechnology, St. Xavier's College, 30 Park Street, Kolkata 700016, India.
authors.forename: Sanjay
authors.initials: S
authors.lastname: Dey
authors.affiliation: High Pressure &Synchrotron Radiation Physics Division, Bhabha Atomic Research Centre, Trombay, Mumbai 400085, India.
authors.forename: Biplab
authors.initials: B
authors.lastname: Ghosh
authors.affiliation: Department of Biotechnology, St. Xavier's College, 30 Park Street, Kolkata 700016, India.
authors.forename: Maitree
authors.initials: M
authors.lastname: Biswas
authors.affiliation: Department of Biotechnology, St. Xavier's College, 30 Park Street, Kolkata 700016, India.
authors.forename: Jhimli
authors.initials: J
authors.lastname: Dasgupta
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004758:Enterobactin / Q000378:metabolism*; D006418:Heme / Q000378:metabolism; D006877:Hydroxamic Acids / Q000378:metabolism*; D008024:Ligands; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009154:Mutation; D033902:Periplasmic Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D014734:Vibrio cholerae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D006877:Hydroxamic Acids; D008024:Ligands; D033902:Periplasmic Binding Proteins; D004758:Enterobactin; D006418:Heme