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Structure and Energetics of Allosteric Regulation of HCN2 Ion Channels by Cyclic Nucleotides.

DeBerg HA, Brzovic PS, Flynn GE, Zagotta WN, Stoll S - J. Biol. Chem. (2015)

Bottom Line: Binding of cyclic nucleotides increases the rate and extent of channel activation and shifts it to less hyperpolarized voltages.We probed the allosteric mechanism of different cyclic nucleotides on the CNBD and on channel gating.We explain these results with a model where different allosteric mechanisms in the CNBD all converge to have the same effect on the C-linker and render all three cyclic nucleotides similarly potent activators of the channel.

View Article: PubMed Central - PubMed

Affiliation: From the Departments of Chemistry, Physiology and Biophysics, and.

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Chemical shift perturbations of HCN2-CNBD. Chemical shift differences between apo and 200 μm cAMP (red), apo and 200 μm cGMP (green), and apo and 500 μm cCMP (cyan) bound HCN2-CNBD. Resonances that could not be unambiguously assigned after titration of cyclic nucleotides are indicated by gray shaded bars.
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Figure 4: Chemical shift perturbations of HCN2-CNBD. Chemical shift differences between apo and 200 μm cAMP (red), apo and 200 μm cGMP (green), and apo and 500 μm cCMP (cyan) bound HCN2-CNBD. Resonances that could not be unambiguously assigned after titration of cyclic nucleotides are indicated by gray shaded bars.

Mentions: The magnitudes of the chemical shift perturbations for the assigned resonances are plotted in Fig. 4. Residues with chemical shifts that disappeared or could not be tracked upon addition of cyclic nucleotide are shown in gray. A comparison of the chemical shift perturbations induced by binding cGMP or cCMP relative to chemical shift perturbations induced by binding cAMP is shown in supplemental Fig. S1. If the CNBD were to sample only two conformations, the chemical shift perturbations of cAMP, cGMP, and cCMP would be expected to be collinear. However, a comparison of chemical shift perturbations indicates that they are not collinear (Figs. 3, B and C, and supplemental Fig. S1). In contrast to these findings, a previous study with the HCN4 isoform found that the cCMP and cAMP chemical shift perturbations fell largely along a line (32). Differences in the HCN isoform used in these experiments may be the cause of this discrepancy. Our results indicate that the HCN2 CNBD is not limited to sampling two conformations corresponding to cyclic nucleotide bound and unbound, but rather operates in a more complex conformational landscape, adopting different conformations depending on which cyclic nucleotide species is bound. The nature of the structural differences between these conformations is unknown at this point.


Structure and Energetics of Allosteric Regulation of HCN2 Ion Channels by Cyclic Nucleotides.

DeBerg HA, Brzovic PS, Flynn GE, Zagotta WN, Stoll S - J. Biol. Chem. (2015)

Chemical shift perturbations of HCN2-CNBD. Chemical shift differences between apo and 200 μm cAMP (red), apo and 200 μm cGMP (green), and apo and 500 μm cCMP (cyan) bound HCN2-CNBD. Resonances that could not be unambiguously assigned after titration of cyclic nucleotides are indicated by gray shaded bars.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4697172&req=5

Figure 4: Chemical shift perturbations of HCN2-CNBD. Chemical shift differences between apo and 200 μm cAMP (red), apo and 200 μm cGMP (green), and apo and 500 μm cCMP (cyan) bound HCN2-CNBD. Resonances that could not be unambiguously assigned after titration of cyclic nucleotides are indicated by gray shaded bars.
Mentions: The magnitudes of the chemical shift perturbations for the assigned resonances are plotted in Fig. 4. Residues with chemical shifts that disappeared or could not be tracked upon addition of cyclic nucleotide are shown in gray. A comparison of the chemical shift perturbations induced by binding cGMP or cCMP relative to chemical shift perturbations induced by binding cAMP is shown in supplemental Fig. S1. If the CNBD were to sample only two conformations, the chemical shift perturbations of cAMP, cGMP, and cCMP would be expected to be collinear. However, a comparison of chemical shift perturbations indicates that they are not collinear (Figs. 3, B and C, and supplemental Fig. S1). In contrast to these findings, a previous study with the HCN4 isoform found that the cCMP and cAMP chemical shift perturbations fell largely along a line (32). Differences in the HCN isoform used in these experiments may be the cause of this discrepancy. Our results indicate that the HCN2 CNBD is not limited to sampling two conformations corresponding to cyclic nucleotide bound and unbound, but rather operates in a more complex conformational landscape, adopting different conformations depending on which cyclic nucleotide species is bound. The nature of the structural differences between these conformations is unknown at this point.

Bottom Line: Binding of cyclic nucleotides increases the rate and extent of channel activation and shifts it to less hyperpolarized voltages.We probed the allosteric mechanism of different cyclic nucleotides on the CNBD and on channel gating.We explain these results with a model where different allosteric mechanisms in the CNBD all converge to have the same effect on the C-linker and render all three cyclic nucleotides similarly potent activators of the channel.

View Article: PubMed Central - PubMed

Affiliation: From the Departments of Chemistry, Physiology and Biophysics, and.

Show MeSH