title: Dynamics and Energy Contributions for Transport of Unfolded Pertactin through a Protein Nanopore.
abstract: To evaluate the physical parameters governing translocation of an unfolded protein across a lipid bilayer, we studied protein transport through aerolysin, a passive protein channel, at the single-molecule level. The protein model used was the passenger domain of pertactin, an autotransporter virulence protein. Transport of pertactin through the aerolysin nanopore was detected as transient partial current blockades as the unfolded protein partially occluded the aerolysin channel. We compared the dynamics of entry and transport for unfolded pertactin and a covalent end-to-end dimer of the same protein. For both the monomer and the dimer, the event frequency of current blockades increased exponentially with the applied voltage, while the duration of each event decreased exponentially as a function of the electrical potential. The blockade time was twice as long for the dimer as for the monomer. The calculated activation free energy includes a main enthalpic component that we attribute to electrostatic interactions between pertactin and the aerolysin nanopore (despite the low Debye length), plus an entropic component due to confinement of the unfolded chain within the narrow pore. Comparing our experimental results to previous studies and theory suggests that unfolded proteins cross the membrane by passing through the nanopore in a somewhat compact conformation according to the "blob" model of Daoud and de Gennes.
authors: Department of Chemistry & Biochemistry, University of Notre Dame , Notre Dame, Indiana 46556, United States.
authors: Benjamin
authors: B
authors: Cressiot
authors: Department of Chemistry & Biochemistry, University of Notre Dame , Notre Dame, Indiana 46556, United States.
authors: Esther
authors: E
authors: Braselmann
authors: LAMBE UMR 8587 CNRS, University of Cergy-Pontoise , Cergy-Pontoise, France.
authors: Abdelghani
authors: A
authors: Oukhaled
authors: Department of Biochemistry, University of Iowa , Iowa City, Iowa 52242, United States.
authors: Adrian H
authors: AH
authors: Elcock
authors: LAMBE UMR 8587 CNRS, University of Évry-Val-d'Essonne , Évry, France.
authors: Juan
authors: J
authors: Pelta
authors: Department of Chemistry & Biochemistry, University of Notre Dame , Notre Dame, Indiana 46556, United States.|Department of Chemical & Biomolecular Engineering, University of Notre Dame , Notre Dame, Indiana 46556, United States.
authors: Patricia L
authors: PL
authors: Clark
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D058608:Nanopores; D036103:Nanotechnology; D011489:Protein Denaturation; D021381:Protein Transport; D058767:Protein Unfolding; D010566:Virulence Factors, Bordetella / Q000737:chemistry* / Q000378:metabolism
chemical_list: D001425:Bacterial Outer Membrane Proteins; D008051:Lipid Bilayers; D010566:Virulence Factors, Bordetella; C068929:pertactin
keywords: autotransporters; electrical detection; protein nanopore; secretion; single molecule; translocation; unfolded protein