Abstract
PapC ushers are outer-membrane proteins enabling assembly and secretion of P pili in uropathogenic E. coli. Their translocation domain is a large β-barrel occluded by a plug domain, which is displaced to allow the translocation of pilus subunits across the membrane. Previous studies suggested that this gating mechanism is controlled by a β-hairpin and an α-helix. To investigate the role of these elements in allosteric signal communication, we developed a method combining evolutionary and molecular dynamics studies of the native translocation domain and mutants lacking the β-hairpin and/or the α-helix. Analysis of a hybrid residue interaction network suggests distinct regions (residue 'communities') within the translocation domain (especially around β12-β14) linking these elements, thereby modulating PapC gating. Antibiotic sensitivity and electrophysiology experiments on a set of alanine-substitution mutants confirmed functional roles for four of these communities. This study illuminates the gating mechanism of PapC ushers and its importance in maintaining outer-membrane permeability.
Keywords:
E. coli; biophysics; dynamics; evolution; outer membrane protein; structural biology; structure.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Alanine / chemistry
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Alanine / metabolism
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Allosteric Regulation
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Amino Acid Motifs
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Amino Acid Substitution
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Anti-Bacterial Agents / pharmacology
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Cell Membrane Permeability
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Erythromycin / pharmacology
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Escherichia coli Proteins / chemistry*
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Escherichia coli Proteins / genetics
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Escherichia coli Proteins / metabolism
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Fimbriae, Bacterial / chemistry*
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Fimbriae, Bacterial / drug effects
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Fimbriae, Bacterial / genetics
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Fimbriae, Bacterial / metabolism
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Gene Expression
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Membrane Potentials
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Molecular Dynamics Simulation
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Molecular Sequence Data
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Mutation
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Porins / chemistry*
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Porins / genetics
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Porins / metabolism
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Protein Structure, Secondary
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Protein Structure, Tertiary
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Protein Subunits / chemistry*
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Protein Subunits / genetics
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Protein Subunits / metabolism
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Protein Transport
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Signal Transduction
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Uropathogenic Escherichia coli / chemistry*
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Uropathogenic Escherichia coli / drug effects
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Uropathogenic Escherichia coli / genetics
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Uropathogenic Escherichia coli / metabolism
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Vancomycin / pharmacology
Substances
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Anti-Bacterial Agents
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Escherichia coli Proteins
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Porins
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Protein Subunits
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atpG protein, E coli
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Erythromycin
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Vancomycin
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Alanine