Inward rectifier potassium channel Kir-2.3 is inhibited by internal sulfhydryl modification

REGIONS of the hippocampal inward rectifier potassium channel Kir 2.3 that contact the aqueous environment were investigated by identification of native cysteine residues that confer sulfhydryl reagent sensitivity to the channel conductance. Kir 2.3 currents were inhibited by N-ethylmaleimide (NEM), whereas currents of Kir 2.1 were unaffected. The reactive residues were identified as Kir 2.3 Cys28 and Cys50 using chimeric constructs and mutagenesis. These sites were not accessible to p-chloromercuriphenylsulfonate (pCMPS) applied extracellularly. However, both Cys28 and Cys50 were accessible to 2-(trimethylammoniumethyl) methanethiosulfonate (MTSET) applied to the intracellular surface of the membrane. These studies demonstrate that Cys28 and Cys50 lie in a cytoplasmic aqueous accessible region of the channel, and suggest that the channel N-terminus is a key constituent of the internal vestibule of the pore and/or modulates channel gating. NeuroReport 10:3277-3282 (C) 1999 Lippincott Williams & Wilkins.