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Effect of endosomal acidification on small ion transport through the anthrax toxin PA63 channel.

ABSTRACT: Tight regulation of pH is critical for the structure and function of cells and organelles. The pH environment changes dramatically along the endocytic pathway, an internalization transport process that is 'hijacked' by many intracellularly active bacterial exotoxins, including the anthrax toxin. Here, we investigate the role of pH on single-channel properties of the anthrax toxin protective antigen (PA63 ). Using conductance and current noise analysis, blocker binding, ion selectivity, and poly(ethylene glycol) partitioning measurements, we show that the channel exists in two different open states ('maximum' and 'main') at pH ? 5.5, while only a maximum conductance state is detected at pH < 5.5. We describe two substantially distinct patterns of PA63 conductance dependence on KCl concentration uncovered at pH 6.5 and 4.5.

SUBMITTER: Kalu N

PROVIDER: S-EPMC5683923 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Publications

Effect of endosomal acidification on small ion transport through the anthrax toxin PA<sub>63</sub> channel.

Kalu Nnanya N Alcaraz Antonio A Yamini Goli G Momben Abolfath Sanaz S Lucas Laura L Kenney Clare C Aguilella Vicente M VM Nestorovich Ekaterina M EM

FEBS letters 20171019 21

Tight regulation of pH is critical for the structure and function of cells and organelles. The pH environment changes dramatically along the endocytic pathway, an internalization transport process that is 'hijacked' by many intracellularly active bacterial exotoxins, including the anthrax toxin. Here, we investigate the role of pH on single-channel properties of the anthrax toxin protective antigen (PA<sub>63</sub> ). Using conductance and current noise analysis, blocker binding, ion selectivity ...[more]

PMID: 28963849

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Project description:Obstructing conductive pathways of the channel-forming toxins with targeted blockers is a promising drug design approach. Nearly all tested positively charged ligands have been shown to reversibly block the cation-selective channel-forming protective antigen (PA63) component of the binary anthrax toxin. The cationic ligands with more hydrophobic surfaces, particularly those carrying aromatic moieties, inhibited PA63 more effectively. To understand the physical basis of PA63 selectivity for a particular ligand, detailed information is required on how the blocker structural elements (e.g., positively charged and aromatic groups) influence the molecular kinetics of the blocker/channel binding reactions. In this study, we address this problem using the high-resolution single-channel planar lipid bilayer technique. Several structurally distinct cationic blockers, namely per-6-S-(3-amino) propyl-?-cyclodextrin, per-6-S-(3-aminomethyl) benzyl-?-cyclodextrin, per-6-S-(3-aminomethyl) benzyl-?-cyclodextrin, per-6-S-(3-aminomethyl) benzyl-?-cyclodextrin, methyltriphenylphosphonium ion, and G0 polyamidoamine dendrimer are tested for their ability to inhibit the heptameric and octameric PA63 variants and PA63F427A mutant. The F427 residues form a hydrophobic constriction region inside the channel, known as the "?-clamp." We show that the cationic blockers interact with PA63 through a combination of forces. Analysis of the binding reaction kinetics suggests the involvement of cation-?, Coulomb, and salt-concentration-independent ?-? or hydrophobic interactions in the cationic cyclodextrin binding. It is possible that these blockers bind to the ?-clamp and are also stabilized by the Coulomb interactions of their terminal amino groups with the water-exposed negatively charged channel residues. In PA63F427A, only the suggested Coulomb component of the cyclodextrin interaction remains. Methyltriphenylphosphonium ion and G0 polyamidoamine dendrimer, despite being positively charged, interact primarily with the ?-clamp. We also show that seven- and eightfold symmetric cyclodextrins effectively block the heptameric and octameric forms of PA63 interchangeably, adding flexibility to the earlier formulated blocker/target symmetry match requirement.

Dataset Information

Effect of endosomal acidification on small ion transport through the anthrax toxin PA63 channel.

Publications

Effect of endosomal acidification on small ion transport through the anthrax toxin PA<sub>63</sub> channel.

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