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Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block

The internal vestibule of large-conductance Ca(2+) voltage-activated K(+) (BK) channels contains a ring of eight negative charges not present in K(+) channels of lower conductance (Glu386 and Glu389 in hSlo) that modulates channel conductance through an electrostatic mechanism (Brelidze, T.I., X. Ni...

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Autores principales: Carvacho, Ingrid, Gonzalez, Wendy, Torres, Yolima P., Brauchi, Sebastian, Alvarez, Osvaldo, Gonzalez-Nilo, Fernando D., Latorre, Ramon
Formato: Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2008
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2213566/
https://www.ncbi.nlm.nih.gov/pubmed/18227273
http://dx.doi.org/10.1085/jgp.200709862
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author Carvacho, Ingrid
Gonzalez, Wendy
Torres, Yolima P.
Brauchi, Sebastian
Alvarez, Osvaldo
Gonzalez-Nilo, Fernando D.
Latorre, Ramon
author_facet Carvacho, Ingrid
Gonzalez, Wendy
Torres, Yolima P.
Brauchi, Sebastian
Alvarez, Osvaldo
Gonzalez-Nilo, Fernando D.
Latorre, Ramon
author_sort Carvacho, Ingrid
collection PubMed
description The internal vestibule of large-conductance Ca(2+) voltage-activated K(+) (BK) channels contains a ring of eight negative charges not present in K(+) channels of lower conductance (Glu386 and Glu389 in hSlo) that modulates channel conductance through an electrostatic mechanism (Brelidze, T.I., X. Niu, and K.L. Magleby. 2003. Proc. Natl. Acad. Sci. USA. 100:9017–9022). In BK channels there are also two acidic amino acid residues in an extracellular loop (Asp326 and Glu329 in hSlo). To determine the electrostatic influence of these charges on channel conductance, we expressed wild-type BK channels and mutants E386N/E389N, D326N, E329Q, and D326N/E329Q channels on Xenopus laevis oocytes, and measured the expressed currents under patch clamp. Contribution of E329 to the conductance is negligible and single channel conductance of D326N/E329Q channels measured at 0 mV in symmetrical 110 mM K(+) was 18% lower than the control. Current–voltage curves displayed weak outward rectification for D326N and the double mutant. The conductance differences between the mutants and wild-type BK were caused by an electrostatic effect since they were enhanced at low K(+) (30 mM) and vanished at high K(+) (1 M K(+)). We determine the electrostatic potential change, Δφ, caused by the charge neutralization using TEA(+) block for the extracellular charges and Ba(2+) for intracellular charges. We measured 13 ± 2 mV for Δφ at the TEA(+) site when turning off the extracellular charges, and 17 ± 2 mV for the Δφ at the Ba(2+) site when the intracellular charges were turned off. To understand the electrostatic effect of charge neutralizations, we determined Δφ using a BK channel molecular model embedded in a lipid bilayer and solving the Poisson-Boltzmann equation. The model explains the experimental results adequately and, in particular, gives an economical explanation to the differential effect on the conductance of the neutralization of charges D326 and E329.
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spelling pubmed-22135662008-08-01 Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block Carvacho, Ingrid Gonzalez, Wendy Torres, Yolima P. Brauchi, Sebastian Alvarez, Osvaldo Gonzalez-Nilo, Fernando D. Latorre, Ramon J Gen Physiol Articles The internal vestibule of large-conductance Ca(2+) voltage-activated K(+) (BK) channels contains a ring of eight negative charges not present in K(+) channels of lower conductance (Glu386 and Glu389 in hSlo) that modulates channel conductance through an electrostatic mechanism (Brelidze, T.I., X. Niu, and K.L. Magleby. 2003. Proc. Natl. Acad. Sci. USA. 100:9017–9022). In BK channels there are also two acidic amino acid residues in an extracellular loop (Asp326 and Glu329 in hSlo). To determine the electrostatic influence of these charges on channel conductance, we expressed wild-type BK channels and mutants E386N/E389N, D326N, E329Q, and D326N/E329Q channels on Xenopus laevis oocytes, and measured the expressed currents under patch clamp. Contribution of E329 to the conductance is negligible and single channel conductance of D326N/E329Q channels measured at 0 mV in symmetrical 110 mM K(+) was 18% lower than the control. Current–voltage curves displayed weak outward rectification for D326N and the double mutant. The conductance differences between the mutants and wild-type BK were caused by an electrostatic effect since they were enhanced at low K(+) (30 mM) and vanished at high K(+) (1 M K(+)). We determine the electrostatic potential change, Δφ, caused by the charge neutralization using TEA(+) block for the extracellular charges and Ba(2+) for intracellular charges. We measured 13 ± 2 mV for Δφ at the TEA(+) site when turning off the extracellular charges, and 17 ± 2 mV for the Δφ at the Ba(2+) site when the intracellular charges were turned off. To understand the electrostatic effect of charge neutralizations, we determined Δφ using a BK channel molecular model embedded in a lipid bilayer and solving the Poisson-Boltzmann equation. The model explains the experimental results adequately and, in particular, gives an economical explanation to the differential effect on the conductance of the neutralization of charges D326 and E329. The Rockefeller University Press 2008-02 /pmc/articles/PMC2213566/ /pubmed/18227273 http://dx.doi.org/10.1085/jgp.200709862 Text en Copyright © 2008, The Rockefeller University Press This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Articles
Carvacho, Ingrid
Gonzalez, Wendy
Torres, Yolima P.
Brauchi, Sebastian
Alvarez, Osvaldo
Gonzalez-Nilo, Fernando D.
Latorre, Ramon
Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block
title Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block
title_full Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block
title_fullStr Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block
title_full_unstemmed Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block
title_short Intrinsic Electrostatic Potential in the BK Channel Pore: Role in Determining Single Channel Conductance and Block
title_sort intrinsic electrostatic potential in the bk channel pore: role in determining single channel conductance and block
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2213566/
https://www.ncbi.nlm.nih.gov/pubmed/18227273
http://dx.doi.org/10.1085/jgp.200709862
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