An engineered cysteine in the external mouth of a K+ channel allows inactivation to be modulated by metal binding.
Substitution of a cysteine in the extracellular mouth of the pore of the Shaker-delta K+ channel permits allosteric inhibition of the channel by Zn2+ or Cd2+ ions at micromolar concentrations. Cd2+ binds weakly to the open state but drives the channel into the slow (C-type) inactivated state, which has a Kd for Cd2+ of approximately 0.2 microM. There is a 45,000-fold increase in affinity when the channel changes from open to inactivated. These results indicate that C-type inactivation involves a structural change in the external mouth of the pore. This structural change is reflected in the T449C mutant as state-dependent metal affinity, which may result either from a change in proximity of the introduced cysteine residues of the four subunits or from a change of the exposure of this residue on the surface of the protein.
Yellen
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Department of Neurobiology, Harvard Medical School, Boston, Massachusetts.
Sodickson
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D
Chen
T Y
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Jurman
M E
ME
D000494:Allosteric Regulation / Q000235:genetics; D000818:Animals; D001665:Binding Sites; D055592:Biophysical Phenomena; D001703:Biophysics; D002104:Cadmium / Q000378:metabolism / Q000494:pharmacology; D002460:Cell Line; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D015202:Protein Engineering; D015032:Zinc / Q000378:metabolism
D026902:Potassium Channel Blockers; D015221:Potassium Channels; D002104:Cadmium; D015032:Zinc; D003545:Cysteine