Co-expression of wild-type and mutant olfactory cyclic nucleotide-gated channels: restoration of the native sensitivity to Ca(2+) and Mg(2+) blockage.
In the pore of homomeric cyclic nucleotide-gated (CNG) channels, Ca(2+) and Mg(2+) bind to a set of glutamate residues, which in the bovine olfactory CNG channel are located at position 340. However, native CNG channels from olfactory sensory neurons are composed by the assembly of three different types of subunits, each having a different residue -- glutamate, aspartate or glycine -- at the position corresponding to the binding site for external Ca(2+) and Mg(2+). We co-expressed the wild-type principal alpha subunit with its mutants E340G and E340D in different combinations in Xenopus laevis oocytes, and measured Ca(2+) and Mg(2+) blockage in excised outside-out membrane patches. The comparison between our results and data from native olfactory CNG channels indicates that the presence of all three residues -- glutamate, aspartate and glycine -- in the different subunits, is necessary to restore the sensitivity to external Ca(2+) and Mg(2+) measured in native channels.
Picco
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Istituto di Cibernetica e Biofisica, Consiglio Nazionale delle Ricerche, Via De Marini 6, 16149 Genova, Italy.
Gavazzo
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Menini
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D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002417:Cattle; D000242:Cyclic AMP / Q000494:pharmacology; D006152:Cyclic GMP / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels; D005260:Female; D015870:Gene Expression / Q000502:physiology; D015640:Ion Channel Gating / Q000187:drug effects* / Q000502:physiology; D007473:Ion Channels / Q000235:genetics* / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology*; D016297:Mutagenesis, Site-Directed / Q000502:physiology; D018034:Olfactory Receptor Neurons / Q000502:physiology; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger / Q000494:pharmacology; D014982:Xenopus laevis
D054815:Cyclic Nucleotide-Gated Cation Channels; D007473:Ion Channels; D012333:RNA, Messenger; D000242:Cyclic AMP; D006152:Cyclic GMP; D008274:Magnesium; D002118:Calcium