title: Interaction of charybdotoxin with permeant ions inside the pore of a K+ channel.
abstract: Charybdotoxin (CTX) blocks high conductance Ca(2+)-activated K+ channels by binding to a receptor site in the externally facing "mouth." Toxin bound to the channel can be destabilized from its site by K+ entering the channel from the opposite, internal, solution. By analyzing point mutants of CTX expressed in E. coli, assayed with single Ca(2+)-activated K+ channels reconstituted into planar lipid bilayers, we show that a single positively charged residue of the peptide, Lys-27, wholly mediates this interaction of K+ with CTX. If position 27 carries a positively charged residue, internal K+ accelerates the dissociation rate of CTX in a voltage-dependent manner; however, if a neutral Asn or Gln is substituted at this position, the dissociation rate is completely insensitive to either internal K+ or applied voltage. Position 27 is unique in this respect; charge-neutral substitutions made at other positions fail to eliminate the K+ destabilization phenomenon. The results argue that CTX bound to the channel positions Lys-27 physically close to a K(+)-specific binding site on the external end of the conduction pathway and that a K+ ion occupying this site destabilizes CTX via direct electrostatic repulsion with the epsilon-amino group of Lys-27.
authors.affiliation: Howard Hughes Medical Institute, Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254-9110.
authors.forename: C S
authors.initials: CS
authors.lastname: Park
authors.forename: C
authors.initials: C
authors.lastname: Miller
mesh_terms: D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D018999:Charybdotoxin; D004553:Electric Conductivity; D004563:Electrochemistry; D004926:Escherichia coli / Q000378:metabolism; D007700:Kinetics; D008051:Lipid Bilayers / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D016296:Mutagenesis; D011188:Potassium / Q000378:metabolism / Q000494:pharmacology; D015221:Potassium Channels / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012604:Scorpion Venoms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D008051:Lipid Bilayers; D015221:Potassium Channels; D011994:Recombinant Proteins; D012604:Scorpion Venoms; D018999:Charybdotoxin; D008239:Lysine; D011188:Potassium; D002118:Calcium