Tracking the movement of discrete gating charges in a voltage-gated potassium channel

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Abstract

Positively-charged amino acids respond to membrane potential changes to drive voltage sensor movement in voltage-gated ion channels, but determining the displacements of voltage sensor gating charges has proven difficult. We optically tracked the movement of the two most extracellular charged residues (R1, R2) in the Shaker potassium channel voltage sensor using a fluorescent positively-charged bimane derivative (qBBr) that is strongly quenched by tryptophan. By individually mutating residues to tryptophan within the putative pathway of gating charges, we observed that the charge motion during activation is a rotation and a tilted translation that differs between R1 and R2. Tryptophan-induced quenching of qBBr also indicates that a crucial residue of the hydrophobic plug is linked to the Cole-Moore shift through its interaction with R1. Finally, we show that this approach extends to additional voltage-sensing membrane proteins using the Ciona intestinalis voltage sensitive phosphatase (CiVSP) (Murata et al., 2005a).

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All data generated or analysed during this study are included in the manuscript and supporting files. Actual records have been provided for Figures 2,3,4,5,7,8

Article and author information

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Michael F Priest

Biochemistry and Molecular Biology, University of Chicago, Chicago, United States

Competing interests
The authors declare that no competing interests exist.
Elizabeth EL Lee

Biochemistry and Molecular Biology, University of Chicago, Chicago, United States

Competing interests
The authors declare that no competing interests exist.
Francisco Bezanilla

Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, United States

For correspondence
fbezanilla@uchicago.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-6663-7931

Funding

National Institutes of Health (R01-GM030376)

Francisco Bezanilla

National Institutes of Health (F31NS081954)

Michael F Priest

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All of the animals were handled according to approved institutional animal care and use committee (IACUC) protocols of the University of Chicago. The protocol was approved by the Committee on the Ethics of Animal Experiments of the University of Chicago (Permit Number: 71475).

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.