|
|
|
|
|
|
|
||
Original Article |
Richard.horn{at}mail.tju.edu
Voltage-gated ion channels have at least two classes of moving parts, voltage sensors that respond to changes in the transmembrane potential and gates that create or deny permeant ions access to the conduction pathway. To explore the coupling between voltage sensors and gates, we have systematically immobilized each using a bifunctional photoactivatable cross-linker, benzophenone-4-carboxamidocysteine methanethiosulfonate, that can be tethered to cysteines introduced into the channel protein by mutagenesis. To validate the method, we first tested it on the inactivation gate of the sodium channel. The benzophenone-labeled inactivation gate of the sodium channel can be trapped selectively either in an open or closed state by ultraviolet irradiation at either a hyperpolarized or depolarized voltage, respectively. To verify that ultraviolet light can immobilize S4 segments, we examined its relative effects on ionic and gating currents in Shaker potassium channels, labeled at residue 359 at the extracellular end of the S4 segment. As predicted by the tetrameric stoichiometry of these potassium channels, ultraviolet irradiation reduces ionic current by approximately the fourth power of the gating current reduction, suggesting little cooperativity between the movements of individual S4 segments. Photocross-linking occurs preferably at hyperpolarized voltages after labeling residue 359, suggesting that depolarization moves the benzophenone adduct out of a restricted environment. Immobilization of the S4 segment of the second domain of sodium channels prevents channels from opening. By contrast, photocross-linking the S4 segment of the fourth domain of the sodium channel has effects on both activation and inactivation. Our results indicate that specific voltage sensors of the sodium channel play unique roles in gating, and suggest that movement of one voltage sensor, the S4 segment of domain 4, is at least a two-step process, each step coupled to a different gate.
Key Words: cysteine mutagenesis • sodium channel • Shaker potassium channel • benzophenone • S4 segment
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Facebook 
Reddit 
Technorati 
Twitter What's this?
This article has been cited by other articles:
|
|
 |
|
  S. Sakata, T. Kurokawa, M. H. H. Norholm, M. Takagi, Y. Okochi, G. von Heijne, and Y. Okamura Functionality of the voltage-gated proton channel truncated in S4 PNAS, February 2, 2010; 107(5): 2313 - 2318. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. G. DeCaen, V. Yarov-Yarovoy, E. M. Sharp, T. Scheuer, and W. A. Catterall Sequential formation of ion pairs during activation of a sodium channel voltage sensor PNAS, December 29, 2009; 106(52): 22498 - 22503. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. G. Gagnon and F. Bezanilla A single charged voltage sensor is capable of gating the Shaker K+ channel J. Gen. Physiol., May 1, 2009; 133(5): 467 - 483. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Horn Uncooperative Voltage Sensors J. Gen. Physiol., May 1, 2009; 133(5): 463 - 466. [Full Text] [PDF]
|
|
|
|
 |
|
 V. Yarotskyy, G. Gao, B. Z. Peterson, and K. S. Elmslie The Timothy syndrome mutation of cardiac CaV1.2 (L-type) channels: multiple altered gating mechanisms and pharmacological restoration of inactivation J. Physiol., February 1, 2009; 587(3): 551 - 565. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Okamura, Y. Murata, and H. Iwasaki Voltage-sensing phosphatase: actions and potentials J. Physiol., February 1, 2009; 587(3): 513 - 520. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Muroi and B. Chanda Local Anesthetics Disrupt Energetic Coupling between the Voltage-sensing Segments of a Sodium Channel J. Gen. Physiol., December 29, 2008; 133(1): 1 - 15. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. V. Campos, B. Chanda, P. S.L. Beirao, and F. Bezanilla {alpha}-Scorpion Toxin Impairs a Conformational Change that Leads to Fast Inactivation of Muscle Sodium Channels J. Gen. Physiol., July 28, 2008; 132(2): 251 - 263. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Md. I. Hossain, H. Iwasaki, Y. Okochi, M. Chahine, S. Higashijima, K. Nagayama, and Y. Okamura Enzyme Domain Affects the Movement of the Voltage Sensor in Ascidian and Zebrafish Voltage-sensing Phosphatases J. Biol. Chem., June 27, 2008; 283(26): 18248 - 18259. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. V. Campos, B. Chanda, P. S.L. Beirao, and F. Bezanilla {beta}-Scorpion Toxin Modifies Gating Transitions in All Four Voltage Sensors of the Sodium Channel J. Gen. Physiol., August 27, 2007; 130(3): 257 - 268. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. M. Armstrong Na channel inactivation from open and closed states PNAS, November 21, 2006; 103(47): 17991 - 17996. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. T. Sack and R. W. Aldrich Binding of a Gating Modifier Toxin Induces Intersubunit Cooperativity Early in the Shaker K Channel's Activation Pathway J. Gen. Physiol., June 26, 2006; 128(1): 119 - 132. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. Ulbricht Sodium Channel Inactivation: Molecular Determinants and Modulation Physiol Rev, October 1, 2005; 85(4): 1271 - 1301. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Kitaguchi, M. Sukhareva, and K. J. Swartz Stabilizing the Closed S6 Gate in the Shaker K v Channel Through Modification of a Hydrophobic Seal J. Gen. Physiol., September 27, 2004; 124(4): 319 - 332. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Yang, Y. Yan, and F. J. Sigworth Can Shaker Potassium Channels be Locked in the Deactivated State? J. Gen. Physiol., July 26, 2004; 124(2): 163 - 171. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. P. Faillace, R. O. Bernabeu, and J. I. Korenbrot Cellular Processing of Cone Photoreceptor Cyclic GMP-gated Ion Channels: A ROLE FOR THE S4 STRUCTURAL MOTIF J. Biol. Chem., May 21, 2004; 279(21): 22643 - 22653. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. A. Ahern and R. Horn Specificity of Charge-carrying Residues in the Voltage Sensor of Potassium Channels J. Gen. Physiol., February 23, 2004; 123(3): 205 - 216. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Chanda, O. K. Asamoah, and F. Bezanilla Coupling Interactions between Voltage Sensors of the Sodium Channel as Revealed by Site-specific Measurements J. Gen. Physiol., February 23, 2004; 123(3): 217 - 230. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Sukhareva, D. H. Hackos, and K. J. Swartz Constitutive Activation of the Shaker Kv Channel J. Gen. Physiol., October 27, 2003; 122(5): 541 - 556. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. R Groome, E. Fujimoto, and P. C Ruben Negative charges in the DIII-DIV linker of human skeletal muscle Na+ channels regulate deactivation gating J. Physiol., April 1, 2003; 548(1): 85 - 96. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Chanda and F. Bezanilla Tracking Voltage-dependent Conformational Changes in Skeletal Muscle Sodium Channel during Activation J. Gen. Physiol., October 29, 2002; 120(5): 629 - 645. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Lu, A. M. Klem, and Y. Ramu Coupling between Voltage Sensors and Activation Gate in Voltage-gated K+ Channels J. Gen. Physiol., October 29, 2002; 120(5): 663 - 676. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Horn Coupled Movements in Voltage-gated Ion Channels J. Gen. Physiol., September 30, 2002; 120(4): 449 - 453. [Full Text] [PDF]
|
|
|
|
|
|
F. Bezanilla Voltage Sensor Movements J. Gen. Physiol., September 30, 2002; 120(4): 465 - 473. [Full Text] [PDF]
|
|
|
|
|
|
T. P. Nguyen and R. Horn Movement and Crevices Around a Sodium Channel S3 Segment J. Gen. Physiol., August 26, 2002; 120(3): 419 - 436. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. K Alekov, M. M. Rahman, N. Mitrovic, F. Lehmann-Horn, and H. Lerche A sodium channel mutation causing epilepsy in man exhibits subtle defects in fast inactivation and activation in vitro J. Physiol., December 15, 2000; 529(3): 533 - 539. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Hilber, W. Sandtner, O. Kudlacek, I. W. Glaaser, E. Weisz, J. W. Kyle, R. J. French, H. A. Fozzard, S. C. Dudley, and H. Todt The Selectivity Filter of the Voltage-gated Sodium Channel Is Involved in Channel Activation J. Biol. Chem., July 20, 2001; 276(30): 27831 - 27839. [Abstract] [Full Text] [PDF]
|
|
|
|
