|
|
|
|
|
|
|
||
Article |
Cyclic nucleotide–gated channels contain four subunits, each with a binding site for cGMP or cAMP in the cytoplasmic COOH-terminal domain. Previous studies of the kinetic mechanism of activation have been hampered by the complication that ligands are continuously binding and unbinding at each of these sites. Thus, even at the single channel level, it has been difficult to distinguish changes in behavior that arise from a channel with a fixed number of ligands bound from those that occur upon the binding and unbinding of ligands. For example, it is often assumed that complex behaviors like multiple conductance levels and bursting occur only as a consequence of changes in the number of bound ligands. We have overcome these ambiguities by covalently tethering one ligand at a time to single rod cyclic nucleotide–gated channels (Ruiz, ML., and J.W. Karpen. 1997. Nature. 389:389–392). We find that with a fixed number of ligands locked in place the channel freely moves between three conductance states and undergoes bursting behavior. Furthermore, a thorough kinetic analysis of channels locked in doubly, triply, and fully liganded states reveals more than one kinetically distinguishable state at each conductance level. Thus, even when the channel contains a fixed number of bound ligands, it can assume at least nine distinct states. Such complex behavior is inconsistent with simple concerted or sequential allosteric models. The data at each level of liganding can be successfully described by the same connected state model (with different rate constants), suggesting that the channel undergoes the same set of conformational changes regardless of the number of bound ligands. A general allosteric model, which postulates one conformational change per subunit in both the absence and presence of ligand, comes close to providing enough kinetically distinct states. We propose an extension of this model, in which more than one conformational change per subunit can occur during the process of channel activation.
Key Words: allosteric proteins • ligand-gated ion channels • photoaffinity labeling • patch clamp • retinal rod photoreceptors
Abbreviations: APT, 8-p-azidophenacylthio; CNG channel, cyclic nucleotide–gated channel; KNF, Koshland-Nemethy-Filmer; MWC, Monod-Wyman-Changeux
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Facebook 
Reddit 
Technorati 
Twitter What's this?
This article has been cited by other articles:
|
|
 |
|
  W. Bonigk, A. Loogen, R. Seifert, N. Kashikar, C. Klemm, E. Krause, V. Hagen, E. Kremmer, T. Strunker, and U. B. Kaupp An Atypical CNG Channel Activated by a Single cGMP Molecule Controls Sperm Chemotaxis Sci. Signal., October 27, 2009; 2(94): ra68 - ra68. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. E. Contreras and M. Holmgren Access of Quaternary Ammonium Blockers to the Internal Pore of Cyclic Nucleotide-gated Channels: Implications for the Location of the Gate J. Gen. Physiol., April 24, 2006; 127(5): 481 - 494. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Hallermann, S. Heckmann, J. Dudel, and M. Heckmann Short openings in high resolution single channel recordings of mouse nicotinic receptors J. Physiol., March 15, 2005; 563(3): 645 - 662. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Kusch, V. Nache, and K. Benndorf Effects of permeating ions and cGMP on gating and conductance of rod-type cyclic nucleotide-gated (CNGA1) channels J. Physiol., November 1, 2004; 560(3): 605 - 616. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Raghavachari and J. E. Lisman Properties of Quantal Transmission at CA1 Synapses J Neurophysiol, October 1, 2004; 92(4): 2456 - 2467. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Q. Nai, J. M. McIntosh, and J. F. Margiotta Relating Neuronal Nicotinic Acetylcholine Receptor Subtypes Defined by Subunit Composition and Channel Function Mol. Pharmacol., February 1, 2003; 63(2): 311 - 324. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Rosenbaum, L. D. Islas, A. E. Carlson, and S. E. Gordon Dequalinium: A Novel, High-affinity Blocker of CNGA1 Channels J. Gen. Physiol., December 30, 2002; 121(1): 37 - 47. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 U. B. Kaupp and R. Seifert Cyclic Nucleotide-Gated Ion Channels Physiol Rev, July 1, 2002; 82(3): 769 - 824. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Bowie and G. D. Lange Functional Stoichiometry of Glutamate Receptor Desensitization J. Neurosci., May 1, 2002; 22(9): 3392 - 3403. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. R Laver, G. K E Lenz, and G. D Lamb Regulation of the calcium release channel from rabbit skeletal muscle by the nucleotides ATP, AMP, IMP and adenosine J. Physiol., December 15, 2001; 537(3): 763 - 778. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Robert, S. N. Irizarry, T. E. Hughes, and J. R. Howe Subunit Interactions and AMPA Receptor Desensitization J. Neurosci., August 1, 2001; 21(15): 5574 - 5586. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Molokanova and R. H. Kramer Mechanism of Inhibition of Cyclic Nucleotide-Gated Channel by Protein Tyrosine Kinase Probed with Genistein J. Gen. Physiol., March 1, 2001; 117(3): 219 - 234. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. R. Middendorf and R. W. Aldrich Effects of Ultraviolet Modification on the Gating Energetics of Cyclic Nucleotide-Gated Channels J. Gen. Physiol., August 1, 2000; 116(2): 253 - 282. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. L. Anthony, H. L. Brooks, D. Boassa, S. Leonov, G. M. Yanochko, J. W. Regan, and A. J. Yool Cloned Human Aquaporin-1 Is a Cyclic GMP-Gated Ion Channel Mol. Pharmacol., March 1, 2000; 57(3): 576 - 588. [Abstract] [Full Text]
|
|
|
|
