Journal Article

Mechanism of ligand-gated potassium efflux in bacterial pathogens

Roosild, Tarmo P., Booth, Ian R., Miller, Samantha, Pliotas, Christos, Conway, Stuart J., Healy, Jess, Rasmussen, Tim, Castronovo, Samantha, Rees, Douglas C., Bartlett, Wendy

2010

Overview

Gram negative pathogens are protected against toxic electrophilic compounds by glutathione-gated potassium efflux systems (Kef) that modulate cytoplasmic pH. We have elucidated the mechanism of gating through structural and functional analysis of Escherichia coli KefC. The revealed mechanism can explain how subtle chemical differences in glutathione derivatives can produce opposite effects on channel function. Kef channels are regulated by potassium transport and NAD-binding (KTN) domains that sense both reduced glutathione, which inhibits Kef activity, and glutathione adducts that form during electrophile detoxification and activate Kef. We find that reduced glutathione stabilizes an interdomain association between two KTN folds, whereas large adducts sterically disrupt this interaction. F441 is identified as the pivotal residue discriminating between reduced glutathione and its conjugates. We demonstrate a major structural change on the binding of an activating ligand to a KTN-domain protein. Analysis of the regulatory interactions suggests strategies to disrupt pathogen potassium and pH homeostasis.

• Publisher: National Academy of Sciences,National Acad Sciences
• Subject: Adducts / Amino Acid Sequence / Bacterial proteins / Binding sites / Biological Sciences / Biological Transport - drug effects / Channel gating / Crystal structure / Crystallization / Crystals / Detoxification / E coli / Escherichia coli / Escherichia coli - metabolism / Escherichia coli Proteins - chemistry / Escherichia coli Proteins - metabolism / Functional analysis / Glutathione / Glutathione - analogs & derivatives / Glutathione - metabolism / Glutathione - pharmacology / Gram-negative bacteria / Homeostasis / Ion Channel Gating - drug effects / Ion Channel Gating - physiology / Ligands / Models, Molecular / Molecular Sequence Data / Molecular structure / Molecules / Mutant Proteins - chemistry / Mutant Proteins - metabolism / Pathogens / pH effects / Physiological regulation / Potassium / Potassium - metabolism / Potassium Channels - chemistry / Potassium Channels - metabolism / Protein Binding - drug effects / Protein folding / Protein Multimerization - drug effects / Protein Structure, Tertiary / Structure-function relationships / Succinimides - pharmacology / toxicity