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Original Article |
kben{at}mti-n.uni-jena.de
The
subunits of CNG channels of retinal photoreceptors (rod) and olfactory neurons (olf) are proteins that consist of a cytoplasmic NH2 terminus, a transmembrane core region (including the segments S1–S6), and a cytoplasmic COOH terminus. The COOH terminus contains a cyclic nucleotide monophosphate binding domain NBD) that is linked by the C-linker (CL) to the core region. The binding of cyclic nucleotides to the NBD promotes channel opening by an allosteric mechanism. We examined why the sensitivity to cGMP is 22 times higher in olf than in rod by constructing chimeric channels and determining the [cGMP] causing half maximum channel activity (EC50). The characteristic difference in the EC50 value between rod and olf was introduced by the NH2 terminus and the core-CL region, whereas the NBD showed a paradoxical effect. The difference of the free energy difference 
(G) was determined for each of these three regions with all possible combinations of the other two regions. For rod regions with respect to corresponding olf regions, the open channel conformation was destabilized by the NH2 terminus ((G) = –1.0 to –2.0 RT) and the core-CL region ((G) = –2.0 to –2.9 RT), whereas it was stabilized by the NBD ((G) = 0.3 to 1.1 RT). The NH2 terminus deletion mutants of rod and olf differed by (G) of only 0.9 RT, whereas the wild-type channels differed by the much larger value of 3.1 RT. The results show that in rod and olf, the NH2 terminus, the core-CL region, and the NBD differ by characteristic (G) values that do not depend on the specific composition of the other two regions and that the NH2 terminus generates the main portion of (G) between the wild-type channels.
Key Words: ion channel • ligand • cyclic nucleotide • gating
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