Confined Mobility of TonB and FepA in Escherichia coli Membranes

• Published in: PloS one 2016-12, Vol.11 (12), p.e0160862
• Main Authors: Lill, Yoriko, Jordan, Lorne D, Smallwood, Chuck R, Newton, Salete M, Lill, Markus A, Klebba, Phillip E, Ritchie, Ken
• Format: Article
• Language: English
• Subjects: Antibodies, Neutralizing - pharmacology; Bacteria; Bacterial Outer Membrane Proteins - antagonists & inhibitors; Bacterial Outer Membrane Proteins - genetics; Bacterial Outer Membrane Proteins - metabolism; Bacteriology; Biochemistry; Biology and Life Sciences; Biophysics; Carrier Proteins - antagonists & inhibitors; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell Membrane - metabolism; Cell Membrane - ultrastructure; Clonal deletion; Computer simulation; Cytoskeleton; Cytoskeleton - metabolism; Cytoskeleton - ultrastructure; Diffusion; Diffusion coefficient; E coli; Enterobactin; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli - ultrastructure; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Ferric enterobactin; Gene Deletion; Gene Expression Regulation, Bacterial; Kinases; Lateral diffusion; Ligands; Medicine and Health Sciences; Membrane potential; Membrane proteins; Membrane Proteins - genetics; Membrane Proteins - metabolism; Membranes; Mobility; Molecular Dynamics Simulation; Nutrient uptake; Nutrients; Physical Sciences; Physiological aspects; Protein Binding; Protein Transport; Proteins; Receptors, Cell Surface - antagonists & inhibitors; Receptors, Cell Surface - genetics; Receptors, Cell Surface - metabolism; Research and Analysis Methods; Single Molecule Imaging; Time-Lapse Imaging; Transporter
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0160862

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.