Mechanistic signs of double-barreled structure in a fluoride ion channel

Abstract

The Fluc family of F- ion channels protects prokaryotes and lower eukaryotes from the toxicity of environmental F-. In bacteria, these channels are built as dual-topology dimers whereby the two subunits assemble in antiparallel transmembrane orientation. Recent crystal structures suggested that Fluc channels contain two separate ion-conduction pathways, each with two F- binding sites, but no functional correlates of this unusual architecture have been reported. Experiments here fill this gap by examining the consequences of mutating two conserved F-- coordinating phenylalanine residues. Substitution of each phenylalanine specifically extinguishes its associated F- binding site in crystal structures and concomitantly inhibits F- permeation. Functional analysis of concatemeric channels, which permit mutagenic manipulation of individual pores, show that each pore can be separately inactivated without blocking F- conduction through its symmetry-related twin. The results strongly support dual-pathway architecture of Fluc channels.