Key result
The T63S mutation in the selectivity filter of the viral potassium channel Kcv reduced sensitivity to barium block by approximately 100-fold and also conferred resistance to amantadine.
Effect estimate: ~100 fold change
Absolute Event Rate: 818% vs 8.8%
Genetic selection reveals that a single mutation (T63S) in the selectivity filter of the viral potassium channel Kcv confers resistance to both barium and amantadine, mapping their binding to site 4.
Maps blocker sites in viral K+ channels; leaves open translation to human antiviral applications.
BACKGROUND: Understanding the interactions between ion channels and blockers remains an important goal that has implications for delineating the basic mechanisms of ion channel function and for the discovery and development of ion channel directed drugs. METHODOLOGY/PRINCIPAL FINDINGS: We used genetic selection methods to probe the interaction of two ion channel blockers, barium and amantadine, with the miniature viral potassium channel Kcv. Selection for Kcv mutants that were resistant to either blocker identified a mutant bearing multiple changes that was resistant to both. Implementation of a PCR shuffling and backcrossing procedure uncovered that the blocker resistance could be attributed to a single change, T63S, at a position that is likely to form the binding site for the inner ion in the selectivity filter (site 4). A combination of electrophysiological and biochemical assays revealed a distinct difference in the ability of the mutant channel to interact with the blockers. Studies of the analogous mutation in the mammalian inward rectifier Kir2.1 show that the T-->S mutation affects barium block as well as the stability of the conductive state. Comparison of the effects of similar barium resistant mutations in Kcv and Kir2.1 shows that neighboring amino acids in the Kcv selectivity filter affect blocker binding. CONCLUSIONS/SIGNIFICANCE: The data support the idea that permeant ions have an integral role in stabilizing potassium channel structure, suggest that both barium and amantadine act at a similar site, and demonstrate how genetic selections can be used to map blocker binding sites and reveal mechanistic features.
No takes yet. Share an insight, caveat, or question.
Chatelain et al. (2009) studied this question. T63S mutation in Kcv potassium channel vs. Wild-type Kcv channel was evaluated on Barium dissociation constant (Kd) at -80 mV (~100 fold change). The T63S mutation in the selectivity filter of the viral potassium channel Kcv reduced sensitivity to barium block by approximately 100-fold and also conferred resistance to amantadine.
Synapse has enriched 4 closely related papers on similar clinical questions. Consider them for comparative context: