A gating mutation in the internal pore of ASIC1a.
Using a substituted cysteine accessibility scan, we have investigated the structures that form the internal pore of the acid-sensing ion channel 1a. We have identified the amino acid residues Ala-22, Ile-33, and Phe-34 in the amino terminus and Arg-43 in the first transmembrane helix, which when mutated into cysteine, were modified by intracellular application of MTSET, resulting in channel inhibition. The inhibition of the R43C mutant by internal MTSET requires opening of the channel. In addition, binding of Cd2+ ions to R43C slows the channel inactivation. This indicates that the first transmembrane helix undergoes conformational changes during channel inactivation. The effect of Cd2+ on R43C can be obtained with Cd2+ applied at either the extracellular or the intracellular side, indicating that R43C is located in the channel pore. The block of the A22C, I33C, and F34C mutants by MTSET suggests that these residues in the amino terminus of the channel also participate to the internal pore.
Department of Pharmacology and Toxicology, Faculty of Biology and Medicine, Lausanne University, Lausanne CH1005, Switzerland.
Yvan
Y
Pfister
Ivan
I
Gautschi
Armelle-N
AN
Takeda
Miguel
M
van Bemmelen
Stephan
S
Kellenberger
Laurent
L
Schild
D062565:Acid Sensing Ion Channels; D000818:Animals; D002104:Cadmium / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015640:Ion Channel Gating / Q000502:physiology*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008845:Microinjections; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011487:Protein Conformation; D015222:Sodium Channels / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis / Q000378:metabolism
D062565:Acid Sensing Ion Channels; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D015222:Sodium Channels; D002104:Cadmium; D003545:Cysteine