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  • Review Article
  • Published:

Cyclic nucleotide-gated channels: shedding light on the opening of a channel pore

Nature Reviews Neuroscience volume 2, pages 643–651 (2001)Cite this article

Key Points

  • The term 'gating' refers to the allosteric transition that opens and closes the pore of an ion channel. Channel gating has been the focus of intense investigation, but its structural basis remains elusive. The crystal structure of KcsA, a bacterial potassium channel, has provided a framework for new studies of gating in many channel proteins, including cyclic nucleotide-gated (CNG) channels, the focus of this review.

  • The sequence of CNG channels is similar to that of KcsA in the region around the pore domain, having a pore helix, a selectivity filter and an inner helix. Site-directed cysteine substitutions at the presumptive pore helix of CNG1 have provided evidence for the rotation of this helix during gating. Similarly, kinetic analysis and studies with channel blockers have provided indirect evidence for movement of the selectivity filter during gating.

  • In the case of the inner helix, a conformational change in this region also seems to occur during channel gating, as illustrated by the spontaneous formation of disulphide bridges between the inner helices of different CNG subunits when the channel is closed, but not when it is open.

  • The linker between the inner helix and the intracellular cyclic nucleotide-binding domain is crucial for the allosteric coupling between ligand binding and channel opening. It has been found that histidine residues that are present in part of the linker region are capable of coordinating Ni2+ ions between subunits, indicating their spatial proximity. Histidine-substitution experiments show that this region of the linker rotates during gating.

  • On the basis of these and other observations, a new structural model for CNG channel gating is emerging. Opening of the channel involves a clockwise rotation of the distal portion of the linker segment. This rigid body movement unwinds the helical bundle at the bottom of the inner helix, leading to a significant increase in the diameter of the pore. The movement of the inner helix then initiates rearrangements in a gate that is presumably located in the selectivity filter.

Abstract

Few proteins have been described functionally in such detail as ion channels. All ion channels open and close their ion-conducting pores, a process referred to as gating. The recent crystallization of the P-loop-containing channel KcsA has cast channel function in a new light. Results relating to a variety of P-loop-containing channels are converging on a common mechanism in which separation of the inner helices that line the pore results in channel opening. At the same time, differences — some subtle and some perhaps more profound — have emerged between channel types. Here we highlight the evidence for a specific conformational change during the gating of cyclic nucleotide-gated channels, and compare and contrast this evidence to that obtained for other channels.

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Figure 1: Membrane topology for subunits of the two main groups of the P-loop-containing family of ion channels.
Figure 2: Structure of the KcsA channel.
Figure 3: Sequence alignment of the pore helix, selectivity filter and inner helix for several channels of the P-loop-containing family.
Figure 4: State-dependence of modification of amino acids in the pore helix of CNG channels.
Figure 5: Conformational changes in the helix bundle reported by disulphide bond formation at S399C in CNG1.
Figure 6: Ni2+ effects on histidine-substituted channels indicate a rotation of the post-TM segment during gating.
Figure 7: Model of the conformational changes in the helix bundle and post-TM segment of CNG1 channels during gating.

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Authors and Affiliations

  1. Department of Physiology and Biophysics, Howard Hughes Medical Institute, University of Washington, Seattle, 98195, Washington, USA

    Galen E. Flynn, J. P. Johnson Jr & William N. Zagotta

  2. William N. Zagotta

Authors
  1. Galen E. Flynn
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Supplementary information

Model of the conformational changes in the helix bundle and post-TM segment of CNG1 channels during gating.

The movie shows bottom and side views of the model of cyclic nucleotide-gated channel 1 (CNG1) during opening and closing. Histidine substitution at the red positions produced channels that were inhibited by nickel. Histidine substitution at the green positions produced channels that were potentiated by nickel. Position 399 is shown in yellow.

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Related links

DATABASE LINKS

BK

IRK

GIRK

KATP

CNG

X-ray crystal structure of KcsA

KcsA

Shaker

SK

MscL

Herg

KV

FURTHER INFORMATION

SWISS-MODEL server

The ion channel web page

The ligand-gated ion channel database

Families of transport proteins

Glossary

ALLOSTERISM (ALLOSTERY)

The property of a macromolecule by which its function is modified by the binding of an effector to a site other than the binding site of the principal reactant, inducing a conformational shift in the macromolecule.

P LOOP

A conserved structural motif found in many different ion channels, which constitutes part of the channel pore.

2P CHANNELS

Channel proteins that contain two pore-forming domains in each subunit. They constitute the so-called 'KCNK' channel family, and function largely as regulated K+-selective leak channels.

SITE-DIRECTED MUTAGENESIS

The generation of a mutation at a predetermined position in a DNA sequence. The most common method involves the use of a chemically synthesized mutant DNA strand that can hybridize with the target molecule.

SHAKER

A voltage-gated channel, the activation of which leads to the appearance of a transient K+ current. It takes its name from Drosophila with mutations in the gene that encodes this protein. These flies display a violent shaking phenotype when under anaesthesia.

C-TYPE INACTIVATION

Two distinct molecular mechanisms for K+ channel inactivation have been described: N-type, which involves occlusion of the pore by an intracellular domain of the channel, and C-type, which involves a conformational change in the outer pore.

ELECTRON PARAMAGNETIC RESONANCE SPECTROSCOPY

When an atom with an unpaired electron is placed in a magnetic field, the spin of the unpaired electron can align, either in the same direction as the field, or in the opposite direction. EPR is used to measure the absorption of microwave radiation that accompanies the transition between those two states.

PROBE MOBILITY

In an EPR experiment, changes in the mobility of spin-labelled residues are indicative of rearrangements in tertiary or quaternary contacts; positive values indicate increased steric contacts (reduced mobility) and negative values point to increased motional freedom.

PROBE-TO-PROBE DISTANCE

In an EPR experiment, this parameter provides an indication of changes in inter-subunit proximity. In the case of P-loop-containing channels, values lower than 1 indicate that the spin-labelled residues move closer to the axis of symmetry of the channel, whereas larger values point to motion away from the symmetry axis.

CPK MODEL

A space-filling atomic model in which the atoms are represented as spheres, the radii of which are proportional to the van der Waals radius of the atom.

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Flynn, G., Johnson, J. & Zagotta, W. Cyclic nucleotide-gated channels: shedding light on the opening of a channel pore. Nat Rev Neurosci 2, 643–651 (2001). https://doi.org/10.1038/35090015

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