ATP-sensitive potassium channels in adult mouse skeletal muscle: characterization of the ATP-binding site.

Single K+-selective channels were studied in excised inside-out membrane patches from dissociated mouse toe muscle fibers. Channels of 74 pS conductance in symmetrical 160 mM KCl solutions were blocked reversibly by 10 microM internal ATP and thus identified as ATP-sensitive K+ channels. The channels were also blocked reversibly by mM concentrations of internal adenosine, adenine and thymine, but not by cytosine and uracil. The efficacy of the reversible channel blockers was higher when they were present in internal NaCl instead of KCl solutions. An irreversible inhibition of ATP-sensitive K+ channels was observed after application of several sulphydryl-modifying substances in the internal solution: 0.5 mM chloramine-T, 50 mM hydrogen peroxide or 2 mM N-ethylmaleimide (NEM). Large-conductance Ca-activated K+ channels were not affected by these reagents. The presence of 1 mM internal ATP prevents the irreversible inhibition of ATP-sensitive K+ channels by NEM. The results suggest that internal Na+ ions increase the affinity of the ATP-sensitive K+ channel to ATP and to other reversible channel blockers and that a functionally important SH-group is located at or near the ATP-binding site.