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

J Biol Chem. 2016 Jan 1;291(1):371-81. doi: 10.1074/jbc.M115.696450. Epub 2015 Nov 11.


Hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channels play an important role in regulating electrical activity in the heart and brain. They are gated by the binding of cyclic nucleotides to a conserved, intracellular cyclic nucleotide-binding domain (CNBD), which is connected to the channel pore by a C-linker region. 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. Electrophysiology experiments showed that cAMP, cGMP, and cCMP were effective agonists of the channel and produced similar increases in the extent of channel activation. In contrast, electron paramagnetic resonance (EPR) and nuclear magnetic resonance (NMR) on the isolated CNBD indicated that the induced conformational changes and the degrees of stabilization of the active conformation differed for the three cyclic nucleotides. 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.

Keywords: allosteric regulation; cyclic nucleotide; double electron-electron resonance (DEER); electron paramagnetic resonance (EPR); electrophysiology; fluorescence anisotropy; ion channel; nuclear magnetic resonance (NMR).

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation / drug effects
  • Amino Acids / metabolism
  • Animals
  • Anisotropy
  • Electrons
  • Fluorescence
  • Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels / chemistry*
  • Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels / metabolism*
  • Ion Channel Gating / drug effects
  • Magnetic Resonance Spectroscopy
  • Mice
  • Models, Molecular
  • Nucleotides, Cyclic / pharmacology*
  • Potassium Channels / chemistry*
  • Potassium Channels / metabolism*
  • Protein Structure, Tertiary
  • Thermodynamics


  • Amino Acids
  • Hcn2 protein, mouse
  • Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels
  • Nucleotides, Cyclic
  • Potassium Channels

Associated data

  • PDB/3ETQ