Structural implications of weak Ca2+ block in Drosophila cyclic nucleotide-gated channels.
Bottom Line: The selectivity filter of the Drosophila CNG channel is similar to that of most other CNG channels except that it has a threonine at residue 318 instead of a proline.Moreover, mutating the corresponding threonine (T318) to proline in Drosophila CNG channels increased Ca(2+) block by 16 times.These results imply that a simple replacement of a threonine for a proline in Drosophila CNG channels has likely given rise to a distinct selectivity filter conformation that results in weak Ca(2+) block.
Affiliation: Department of Physiology and Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390.Show MeSH
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Mentions: To model the Drosophila CNG channel pore, we replaced the selectivity filter of NaK with that of Drosophila CNG channel and named the construct NaK2CNG-Dm. NaK2CNG-Dm has a filter sequence of 63TVGETPT69, which, in essence, is a P69T mutation of NaK2CNG-E. The structure of NaK2CNG-Dm in complex with K+ was determined to 1.9 Å (Table 1). Although it maintains the same overall structure as NaK (Fig. 2 A), the selectivity filter of NaK2CNG-Dm adopts a novel conformation distinct from that of wild-type NaK, our previous NaK2CNG chimeras, and potassium channel NaK2K (Fig. 2, B and C). The first two filter residues (Thr63 and Val64) of NaK2CNG-Dm form two ion-binding sites (sites 3 and 4) at the internal half of the filter, which are virtually identical to those of other channels. However, the NaK2CNG-Dm filter has a unique external half, where the 65GET filter residues adopt a novel main-chain conformation and generate a deep funnel-shaped entrance filled with layers of ordered water molecules and hydrated K+ ions (Fig. 2 B). This entrance is much deeper and wider than that of the other NaK2CNG mutants such as NaK2CNG-E, to which the detailed structural comparison is made below.
Affiliation: Department of Physiology and Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390.