title: Confined Mobility of TonB and FepA in Escherichia coli Membranes.
abstract: The important process of nutrient uptake in Escherichia coli, in many cases, involves transit of the nutrient through a class of beta-barrel proteins in the outer membrane known as TonB-dependent transporters (TBDTs) and requires interaction with the inner membrane protein TonB. Here we have imaged the mobility of the ferric enterobactin transporter FepA and TonB by tracking them in the membranes of live E. coli with single-molecule resolution at time-scales ranging from milliseconds to seconds. We employed simple simulations to model/analyze the lateral diffusion in the membranes of E.coli, to take into account both the highly curved geometry of the cell and artifactual effects expected due to finite exposure time imaging. We find that both molecules perform confined lateral diffusion in their respective membranes in the absence of ligand with FepA confined to a region [Formula: see text] μm in radius in the outer membrane and TonB confined to a region [Formula: see text] μm in radius in the inner membrane. The diffusion coefficient of these molecules on millisecond time-scales was estimated to be [Formula: see text] μm2/s and [Formula: see text] μm2/s for FepA and TonB, respectively, implying that each molecule is free to diffuse within its domain. Disruption of the inner membrane potential, deletion of ExbB/D from the inner membrane, presence of ligand or antibody to FepA and disruption of the MreB cytoskeleton was all found to further restrict the mobility of both molecules. Results are analyzed in terms of changes in confinement size and interactions between the two proteins.
authors.affiliation: Department of Physics, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Yoriko
authors.initials: Y
authors.lastname: Lill
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Kansas State University, Manhattan, Kansas, United States of America.
authors.forename: Lorne D
authors.initials: LD
authors.lastname: Jordan
authors.affiliation: Department of Chemistry and Biochemistry, University of Oklahoma, Norman, Oklahoma, United States of America.
authors.forename: Chuck R
authors.initials: CR
authors.lastname: Smallwood
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Kansas State University, Manhattan, Kansas, United States of America.
authors.forename: Salete M
authors.initials: SM
authors.lastname: Newton
authors.affiliation: Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Markus A
authors.initials: MA
authors.lastname: Lill
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Kansas State University, Manhattan, Kansas, United States of America.
authors.forename: Phillip E
authors.initials: PE
authors.lastname: Klebba
authors.affiliation: Department of Physics, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Ken
authors.initials: K
authors.lastname: Ritchie
mesh_terms: D057134:Antibodies, Neutralizing / Q000494:pharmacology; D001425:Bacterial Outer Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D003599:Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D004058:Diffusion; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D021381:Protein Transport; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000072760:Single Molecule Imaging; D059008:Time-Lapse Imaging
chemical_list: D057134:Antibodies, Neutralizing; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; C052874:ExbB protein, E coli; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; C029489:enterobactin receptor; C489181:tonB protein, E coli; C060790:exbD protein, E coli; C078473:MreB protein, E coli