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Structural implications of weak Ca2+ block in Drosophila cyclic nucleotide-gated channels.

Lam YL, Zeng W, Derebe MG, Jiang Y - J. Gen. Physiol. (2015)

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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology and Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390.

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Structure of NaK2CNG-Dm. (A) Overall structure of NaK2CNG-Dm, with the front and back subunit removed for clarity. (B) A stereo view of the selectivity filter region boxed in A. 2Fo-Fc (2σ) electron density map is shown in mesh. (C; left), Superimposition of NaK2CNG-Dm (blue), NaK2CNG-E (orange), wild-type NaK (gray), and potassium channel NaK2K (yellow) shows that the four channels differ mainly at the filter region (boxed). (Right) The selectivity filters of NaK2CNG-Dm, NaK2CNG-E, NaK, and NaK2K adopt different conformations at the external half and consequently have varying numbers of ion-binding sites.
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fig2: Structure of NaK2CNG-Dm. (A) Overall structure of NaK2CNG-Dm, with the front and back subunit removed for clarity. (B) A stereo view of the selectivity filter region boxed in A. 2Fo-Fc (2σ) electron density map is shown in mesh. (C; left), Superimposition of NaK2CNG-Dm (blue), NaK2CNG-E (orange), wild-type NaK (gray), and potassium channel NaK2K (yellow) shows that the four channels differ mainly at the filter region (boxed). (Right) The selectivity filters of NaK2CNG-Dm, NaK2CNG-E, NaK, and NaK2K adopt different conformations at the external half and consequently have varying numbers of ion-binding sites.

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.


Structural implications of weak Ca2+ block in Drosophila cyclic nucleotide-gated channels.

Lam YL, Zeng W, Derebe MG, Jiang Y - J. Gen. Physiol. (2015)

Structure of NaK2CNG-Dm. (A) Overall structure of NaK2CNG-Dm, with the front and back subunit removed for clarity. (B) A stereo view of the selectivity filter region boxed in A. 2Fo-Fc (2σ) electron density map is shown in mesh. (C; left), Superimposition of NaK2CNG-Dm (blue), NaK2CNG-E (orange), wild-type NaK (gray), and potassium channel NaK2K (yellow) shows that the four channels differ mainly at the filter region (boxed). (Right) The selectivity filters of NaK2CNG-Dm, NaK2CNG-E, NaK, and NaK2K adopt different conformations at the external half and consequently have varying numbers of ion-binding sites.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4555469&req=5

fig2: Structure of NaK2CNG-Dm. (A) Overall structure of NaK2CNG-Dm, with the front and back subunit removed for clarity. (B) A stereo view of the selectivity filter region boxed in A. 2Fo-Fc (2σ) electron density map is shown in mesh. (C; left), Superimposition of NaK2CNG-Dm (blue), NaK2CNG-E (orange), wild-type NaK (gray), and potassium channel NaK2K (yellow) shows that the four channels differ mainly at the filter region (boxed). (Right) The selectivity filters of NaK2CNG-Dm, NaK2CNG-E, NaK, and NaK2K adopt different conformations at the external half and consequently have varying numbers of ion-binding sites.
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.

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.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology and Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390.

Show MeSH
Related in: MedlinePlus