Modification of cyclic nucleotide-gated ion channels by ultraviolet light.
We irradiated cyclic nucleotide-gated ion channels in situ with ultraviolet light to probe the role of aromatic residues in ion channel function. UV light reduced the current through excised membrane patches from Xenopus oocytes expressing the alpha subunit of bovine retinal cyclic nucleotide-gated channels irreversibly, a result consistent with permanent covalent modification of channel amino acids by UV light. The magnitude of the current reduction depended only on the total photon dose delivered to the patches, and not on the intensity of the exciting light, indicating that the functionally important photochemical modification(s) occurred from an excited state reached by a one-photon absorption process. The wavelength dependence of the channels' UV light sensitivity (the action spectrum) was quantitatively consistent with the absorption spectrum of tryptophan, with a small component at long wavelengths, possibly due to cystine absorption. This spectral analysis suggests that UV light reduced the currents at most wavelengths studied by modifying one or more "target" tryptophans in the channels. Comparison of the channels' action spectrum to the absorption spectrum of tryptophan in various solvents suggests that the UV light targets are in a water-like chemical environment. Experiments on mutant channels indicated that the UV light sensitivity of wild-type channels was not conferred exclusively by any one of the 10 tryptophan residues in a subunit. The similarity in the dose dependences of channel current reduction and tryptophan photolysis in solution suggests that photochemical modification of a small number of tryptophan targets in the channels is sufficient to decrease the currents.
Middendorf
T R
TR
Neurobiology Department, Howard Hughes Medical Institute, School of Medicine, Stanford University, California 94305, USA.
Aldrich
R W
RW
Baylor
D A
DA
D000818:Animals; D002417:Cattle; D003001:Cloning, Molecular; D006152:Cyclic GMP / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels; D004307:Dose-Response Relationship, Radiation; D004553:Electric Conductivity; D015640:Ion Channel Gating / Q000187:drug effects / Q000528:radiation effects*; D007473:Ion Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008564:Membrane Potentials / Q000187:drug effects / Q000528:radiation effects; D008845:Microinjections; D008956:Models, Chemical; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D010777:Photochemistry; D017948:Retinal Rod Photoreceptor Cells / Q000737:chemistry; D012996:Solutions / Q000737:chemistry; D014364:Tryptophan / Q000737:chemistry; D014466:Ultraviolet Rays; D014867:Water / Q000737:chemistry; D014981:Xenopus
D054815:Cyclic Nucleotide-Gated Cation Channels; D007473:Ion Channels; D008024:Ligands; D012996:Solutions; D014867:Water; D014364:Tryptophan; D006152:Cyclic GMP