title: High resolution crystal structures and molecular dynamics studies reveal substrate binding in the porin Omp32.
abstract: The porin Omp32 is the major outer membrane protein of the bacterium Delftia acidovorans. The crystal structures of the strongly anion-selective porin alone and in complex with the substrate malate were solved at 1.5 and 1.45 A resolution, respectively, and revealed a malate-binding motif adjacent to the channel constriction zone. Binding is mediated by interaction with a cluster of two arginine residues and two threonines. This binding site is specific for Omp32 and reflects the physiological adaptation of the organism to organic acids. Structural studies are combined with a 7-ns unbiased molecular dynamics simulation of the trimeric channel in a model membrane. Molecular dynamics trajectories show how malate ions are efficiently captured from the surrounding bulk solution by the electrostatic potential of the channel, translocated to the binding site region, and immobilized in the constriction zone. In accordance with these results, conductance measurements with Omp32 inserted in planar lipid membranes revealed binding of malate. The anion-selective channel Omp32 is the first reported example of a porin with a 16-stranded beta-barrel and proven substrate specificity. This finding suggests a new view on the correlation of porin structure with substrate binding in specific channels.
authors.affiliation: Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried, Germany.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Zachariae
authors.forename: Thomas
authors.initials: T
authors.lastname: Klühspies
authors.forename: Sharmila
authors.initials: S
authors.lastname: De
authors.forename: Harald
authors.initials: H
authors.lastname: Engelhardt
authors.forename: Kornelius
authors.initials: K
authors.lastname: Zeth
mesh_terms: D000838:Anions; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D018360:Crystallography, X-Ray; D020605:Delftia acidovorans / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004583:Electrons; D004594:Electrophysiology; D004926:Escherichia coli / Q000378:metabolism; D006860:Hydrogen Bonding; D007473:Ion Channels; D007477:Ions; D008024:Ligands; D008055:Lipids / Q000737:chemistry; D008293:Malates / Q000737:chemistry; D008958:Models, Molecular; D008968:Molecular Conformation; D018272:Porins / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D021381:Protein Transport; D013379:Substrate Specificity; D013696:Temperature; D013997:Time Factors; D050994:Voltage-Dependent Anion Channels
chemical_list: D000838:Anions; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D007473:Ion Channels; D007477:Ions; D008024:Ligands; D008055:Lipids; D008293:Malates; C115885:Omp32 protein, bacteria; D018272:Porins; D050994:Voltage-Dependent Anion Channels