pH-dependent pore-forming activity of OmpATb from Mycobacterium tuberculosis and characterization of the channel by peptidic dissection
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.
Document Type: Research Article
Affiliations: 1: IBCP, UMR 5086 CNRS, University of Lyon, 69367 Lyon, France. 2: CBS, UMR 5048 CNRS, U 554 INSERM, University of Montpellier I, 34090 Montpellier, France. 3: LDMIM, UMR 5539 CNRS, University of Montpellier II, 34095 Montpellier, France. 4: School of Biological Sciences, University of East Anglia, Norwich NR4 7TJ, UK. 5: The National Institute for Medical Research, Mill Hill, London NW7 1AA, UK.
Publication date: August 1, 2006