|
|
|
|
|
|
|
||
Address correspondence to Henry A. Lester, Division of Biology 156-29, California Institute of Technology, Pasadena, CA 91125. Fax: (626) 564-8709; email: lester{at}caltech.edu
Ion channels of high conductance (>200 pS) are widespread among prokaryotes and eukaryotes. Two examples, the Escherichia coli mechanosensitive ion channels Ec-MscS and Ec-MscL, pass currents of 125–300 pA. To resolve temporal details of conductance transitions, a patch-clamp setup was optimized for low-noise recordings at a time resolution of 3 µs (10–20 times faster than usual). Analyses of the high-resolution recordings confirm that Ec-MscL visits many subconductance states and show that most of the intersubstate transitions occur more slowly than the effective resolution of 3 µs. There is a clear trend toward longer transition times for the larger transitions. In Ec-MscS recordings, the majority of the observed full conductance transitions are also composite. We detected a short-lived (
20 µs) Ec-MscS substate at 2/3 of full conductance; transitions between 2/3 and full conductance did not show fine structure and had a time course limited by the achieved resolution. Opening and closing transitions in MscS are symmetrical and are not preceded or followed by smaller, rapid currents ("anticipations" or "regrets"). Compared with other, lower-conductance channels, these measurements may detect unusually early states in the transitions from fully closed to fully open. Increased temporal resolution at the single-molecule level reveals that some elementary steps of structural transitions are composite and follow several alternative pathways, while others still escape resolution. High-bandwidth, low-noise single-channel measurements may provide details about state transitions in other high-conductance channels; and similar procedures may also be applied to channel- and nanopore-based single-molecule DNA measurements.
Key Words: MscL • MscS • patch clamping • single channel • substate
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  J. S. Santos, S. M. Grigoriev, and M. Montal Molecular Template for a Voltage Sensor in a Novel K+ Channel. III. Functional Reconstitution of a Sensorless Pore Module from a Prokaryotic Kv Channel J. Gen. Physiol., December 1, 2008; 132(6): 651 - 666. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Anishkin, K. Kamaraju, and S. Sukharev Mechanosensitive Channel MscS in the Open State: Modeling of the Transition, Explicit Simulations, and Experimental Measurements of Conductance J. Gen. Physiol., July 1, 2008; 132(1): 67 - 83. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Sotomayor, V. Vasquez, E. Perozo, and K. Schulten Ion Conduction through MscS as Determined by Electrophysiology and Simulation Biophys. J., February 1, 2007; 92(3): 886 - 902. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Tang, G. Cao, X. Chen, J. Yoo, A. Yethiraj, and Q. Cui A Finite Element Framework for Studying the Mechanical Response of Macromolecules: Application to the Gating of the Mechanosensitive Channel MscL Biophys. J., August 15, 2006; 91(4): 1248 - 1263. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Sotomayor, T. A. van der Straaten, U. Ravaioli, and K. Schulten Electrostatic Properties of the Mechanosensitive Channel of Small Conductance MscS Biophys. J., May 15, 2006; 90(10): 3496 - 3510. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Chapman and A. M.J. VanDongen K Channel Subconductance Levels Result from Heteromeric Pore Conformations J. Gen. Physiol., July 25, 2005; 126(2): 87 - 103. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Akitake, A. Anishkin, and S. Sukharev The "Dashpot" Mechanism of Stretch-dependent Gating in MscS J. Gen. Physiol., January 31, 2005; 125(2): 143 - 154. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Anishkin, C.-S. Chiang, and S. Sukharev Gain-of-function Mutations Reveal Expanded Intermediate States and a Sequential Action of Two Gates in MscL J. Gen. Physiol., January 31, 2005; 125(2): 155 - 170. [Abstract] [Full Text] [PDF]
|
|
|
|
