Yeast Trk Proteins Mediate Anion Conduction Via Barrel-Stave Pores

Patch-clamp studies of the potassium-transport proteins Trk1,2 in ascomycete fungi have revealed large chloride efflux currents: at clamp voltages negative to −100 mV, with intracellular chloride concentrations > 10 mM (J. Membr. Biol. 198:177, 2004). Current-voltage analysis of these anion currents, especially in Saccharomyces cerevisiae, led to an in-series two-barrier model for chloride activation: the lower barrier (designated α) being ∼11 kcal/mol and located ∼30% into the membrane from the cytoplasmic surface; and the higher one (β) being ∼14 kcal/mol and located at or near the outer surface. Quantitative adjustments of this model, most importantly in the amplitude of barrier β, efficiently describe almost all current-voltage data for lyotropic anions, with the order of selectivity being I- ≈ Br- > Cl- > SCN- > NO3- at pHo 5.5, and I- ≈ Br- > SCN- > NO3- > Cl-, at pHo 7.5. The kinetic model evokes a hypothetical structure proposed by Durell & Guy (Biophys. J. 77:789, 1999) on the basis of sequence homology with bacterial potassium channels, plus sequence conservation across fungal species. That model posited an intramembrane homotetramer of TRK molecules, arrayed radially around a central cluster of four single helices (TM7) from each monomer. The postulated central cluster would form a hydrophobic pore demarked by a cationic vestibule at the intracellular surface, an intruding ring of tryptophane side-chains near the middle of the membrane, and another ring of phenylalanine side-chains toward the outer surface. The arrangement would be similar to that demonstrated for a variety of (pentameric) ligand-gated ion channels, and suggests a mechanism of voltage-modulated hydrophobic gating to underlie penetration of anions (along with water).