SPATIAL LOCALIZATION OF THE K-BASED STRUCTURE-ANALYSIS( CHANNEL SELECTIVITY FILTER BY MUTANT CYCLE)

The structurally well-characterized scorpion toxin Agitoxin2 inhibits ion permeation through Shaker K+ channels by binding to the external p ore entryway. Scanning mutagenesis identified a set of inhibitor resid ues critical for making energetic contacts with the channel. Using the rmodynamic mutant cycle analysis, we have mapped channel residues rela tive to the known inhibitor structure. This study constrains the posit ion of multiple channel residues within the pore-forming loops; in one stretch, we have been able to map five out of seven contiguous residu es to the inhibitor interaction surface, including those involved in i on selectivity. One interaction in particular, that of K27M on the inh ibitor with Y445F on the channel, is unique in that it depends on the K+ ion concentration. These results reveal a shallow vestibule formed by the pore loops at the K+ channel entryway. The selectivity filter i s located at the center of the vestibule close to (similar to 5 Angstr om) the extracellular solution.