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A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model

Ligand-gated ion channels are oligomers containing several binding sites for the ligands. However, the signal transmission from the ligand binding site to the pore has not yet been fully elucidated for any of these channels. In heteromeric channels, the situation is even more complex than in homomer...

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Autores principales: Benndorf, Klaus, Eick, Thomas, Sattler, Christian, Schmauder, Ralf, Schulz, Eckhard
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Rockefeller University Press 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9066054/
https://www.ncbi.nlm.nih.gov/pubmed/35486087
http://dx.doi.org/10.1085/jgp.202113041
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author Benndorf, Klaus
Eick, Thomas
Sattler, Christian
Schmauder, Ralf
Schulz, Eckhard
author_facet Benndorf, Klaus
Eick, Thomas
Sattler, Christian
Schmauder, Ralf
Schulz, Eckhard
author_sort Benndorf, Klaus
collection PubMed
description Ligand-gated ion channels are oligomers containing several binding sites for the ligands. However, the signal transmission from the ligand binding site to the pore has not yet been fully elucidated for any of these channels. In heteromeric channels, the situation is even more complex than in homomeric channels. Using published data for concatamers of heteromeric cyclic nucleotide–gated channels, we show that, on theoretical grounds, multiple functional parameters of the individual subunits can be determined with high precision. The main components of our strategy are (1) the generation of a defined subunit composition by concatenating multiple subunits, (2) the construction of 16 concatameric channels, which differ in systematically permutated binding sites, (3) the determination of respectively differing concentration–activation relationships, and (4) a complex global fit analysis with corresponding intimately coupled Markovian state models. The amount of constraints in this approach is exceedingly high. Furthermore, we propose a stochastic fit analysis with a scaled unitary start vector of identical elements to avoid any bias arising from a specific start vector. Our approach enabled us to determine 23 free parameters, including 4 equilibrium constants for the closed–open isomerizations, 4 disabling factors for the mutations of the different subunits, and 15 virtual equilibrium-association constants in the context of a 4-D hypercube. From the virtual equilibrium-association constants, we could determine 32 equilibrium-association constants of the subunits at different degrees of ligand binding. Our strategy can be generalized and is therefore adaptable to other ion channels.
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spelling pubmed-90660542022-12-06 A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model Benndorf, Klaus Eick, Thomas Sattler, Christian Schmauder, Ralf Schulz, Eckhard J Gen Physiol Article Ligand-gated ion channels are oligomers containing several binding sites for the ligands. However, the signal transmission from the ligand binding site to the pore has not yet been fully elucidated for any of these channels. In heteromeric channels, the situation is even more complex than in homomeric channels. Using published data for concatamers of heteromeric cyclic nucleotide–gated channels, we show that, on theoretical grounds, multiple functional parameters of the individual subunits can be determined with high precision. The main components of our strategy are (1) the generation of a defined subunit composition by concatenating multiple subunits, (2) the construction of 16 concatameric channels, which differ in systematically permutated binding sites, (3) the determination of respectively differing concentration–activation relationships, and (4) a complex global fit analysis with corresponding intimately coupled Markovian state models. The amount of constraints in this approach is exceedingly high. Furthermore, we propose a stochastic fit analysis with a scaled unitary start vector of identical elements to avoid any bias arising from a specific start vector. Our approach enabled us to determine 23 free parameters, including 4 equilibrium constants for the closed–open isomerizations, 4 disabling factors for the mutations of the different subunits, and 15 virtual equilibrium-association constants in the context of a 4-D hypercube. From the virtual equilibrium-association constants, we could determine 32 equilibrium-association constants of the subunits at different degrees of ligand binding. Our strategy can be generalized and is therefore adaptable to other ion channels. Rockefeller University Press 2022-04-29 /pmc/articles/PMC9066054/ /pubmed/35486087 http://dx.doi.org/10.1085/jgp.202113041 Text en © 2022 Benndorf et al. https://creativecommons.org/licenses/by-nc-sa/4.0/http://www.rupress.org/terms/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 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Article
Benndorf, Klaus
Eick, Thomas
Sattler, Christian
Schmauder, Ralf
Schulz, Eckhard
A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
title A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
title_full A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
title_fullStr A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
title_full_unstemmed A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
title_short A strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
title_sort strategy for determining the equilibrium constants for heteromeric ion channels in a complex model
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9066054/
https://www.ncbi.nlm.nih.gov/pubmed/35486087
http://dx.doi.org/10.1085/jgp.202113041
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