title: pH-dependent pore-forming activity of OmpATb from Mycobacterium tuberculosis and characterization of the channel by peptidic dissection.
abstract: Mycobacteria are characterized by an unusual cell wall that controls nutrient and small hydrophilic compound permeability. Porin-like proteins are necessary to ensure the transport of molecules into the cell. Here, we investigated the pore-forming properties of OmpATb, a porin from Mycobacterium tuberculosis, in lipid bilayers. Multi-channel experiments showed an asymmetric behaviour with channel closures at negative critical voltages (Vc) and a strong decrease in Vc at acidic pH. Single-channel experiments gave conductance values of about 850 +/- 80 pS in 1 M KCl and displayed a weak cationic selectivity in 4-8 pH range. The production and characterization of a series of truncated OmpATb proteins, showed that the central domain (OmpATb73-220) was sufficient to induce the ion channel properties of the native protein in lipid bilayers, i.e. asymmetric insertion, pH-dependent voltage closure, cationic selectivity and similar conductance values in 1 M KCl. Western blot analysis suggests that the presence of OmpATb is only restricted to certain pathogenic species. Therefore, the propensity of channels of native OmpATb to close at low pH may represent an intrinsic property allowing pathogenic mycobacteria to adapt and survive to mildly acidic conditions, such as those encountered within the macrophage phagosome.
authors.affiliation: IBCP, UMR 5086 CNRS, University of Lyon, 69367 Lyon, France.
authors.forename: Virginie
authors.initials: V
authors.lastname: Molle
authors.forename: Nathalie
authors.initials: N
authors.lastname: Saint
authors.forename: Sylvie
authors.initials: S
authors.lastname: Campagna
authors.forename: Laurent
authors.initials: L
authors.lastname: Kremer
authors.forename: Edward
authors.initials: E
authors.lastname: Lea
authors.forename: Philip
authors.initials: P
authors.lastname: Draper
authors.forename: Gérard
authors.initials: G
authors.lastname: Molle
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001703:Biophysics / Q000379:methods; D004594:Electrophysiology / Q000379:methods; D006863:Hydrogen-Ion Concentration; D008051:Lipid Bilayers; D009169:Mycobacterium tuberculosis / Q000472:pathogenicity / Q000502:physiology*; D010446:Peptide Fragments / Q000378:metabolism; D018272:Porins / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D008051:Lipid Bilayers; C113302:OmpATb protein, Mycobacterium tuberculosis; D010446:Peptide Fragments; D018272:Porins; D011994:Recombinant Proteins