Coexpression of alpha and beta subunits of the rod cyclic GMP-gated channel restores native sensitivity to cyclic AMP: role of D604/N1201.
Coexpression of the betawt and alphawt subunits of the bovine rod channel restores two characteristics of the native channels: higher sensitivity to cAMP and potentiation of cGMP-induced currents by low cAMP concentrations. To test whether the increased sensitivity to cAMP is due to the uncharged nature of the asparagine residue (N1201) situated in place of aspartate D604 in the beta subunit as previously suggested (, Neuron. 15:619-625), we compared currents from wild-type (alphawt and alphawt/betawt) and from mutated channels (alphaD604N, alphaD604N/betawt, and alphawt/betaN1201D). The results show that the sensitivity to cAMP and cAMP potentiation is partly but not entirely determined by the charge of residue 1201 in the beta subunit. The D604N mutation in the alpha subunit and, to a lesser extent, coexpression of the betawt subunit with the alphawt subunit reduce the open probability for cGMP compared to that of the alphawt channel. Interpretation of the data with the MWC allosteric model (model of Monod, Wyman, Changeux;, J. Mol. Biol. 12:88-118) suggests that the D604N mutation in the alpha subunits and coassembly of alpha and beta subunits alter the free energy of gating by cAMP more than that of cAMP binding.
Pagès
F
F
Laboratoire de Biophysique Moléculaire et Cellulaire (URA CNRS 520), DBMS, C.E.A.-Grenoble, Grenoble, France.
Ildefonse
M
M
Ragno
M
M
Crouzy
S
S
Bennett
N
N
D019943:Amino Acid Substitution; D000818:Animals; D002417:Cattle; D000242:Cyclic AMP / Q000502:physiology*; D006152:Cyclic GMP / Q000502:physiology*; D054815:Cyclic Nucleotide-Gated Cation Channels; D004110:Diltiazem / Q000494:pharmacology; D015640:Ion Channel Gating; D007473:Ion Channels / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D046911:Macromolecular Substances; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D018408:Patch-Clamp Techniques; D011336:Probability; D017948:Retinal Rod Photoreceptor Cells / Q000502:physiology*
D054815:Cyclic Nucleotide-Gated Cation Channels; D007473:Ion Channels; D046911:Macromolecular Substances; D000242:Cyclic AMP; D004110:Diltiazem; D006152:Cyclic GMP