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In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein

Transient protein-protein and protein-ligand interactions are fundamental components of biological activity. To understand biological activity, not only the structures of the involved proteins are important but also the energetics of the individual steps of a reaction. Here we use in vitro biophysic...

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Autores principales: Niemiec, Moritz S., Weise, Christoph F., Wittung-Stafshede, Pernilla
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3344837/
https://www.ncbi.nlm.nih.gov/pubmed/22574136
http://dx.doi.org/10.1371/journal.pone.0036102
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author Niemiec, Moritz S.
Weise, Christoph F.
Wittung-Stafshede, Pernilla
author_facet Niemiec, Moritz S.
Weise, Christoph F.
Wittung-Stafshede, Pernilla
author_sort Niemiec, Moritz S.
collection PubMed
description Transient protein-protein and protein-ligand interactions are fundamental components of biological activity. To understand biological activity, not only the structures of the involved proteins are important but also the energetics of the individual steps of a reaction. Here we use in vitro biophysical methods to deduce thermodynamic parameters of copper (Cu) transfer from the human copper chaperone Atox1 to the fourth metal-binding domain of the Wilson disease protein (WD4). Atox1 and WD4 have the same fold (ferredoxin-like fold) and Cu-binding site (two surface exposed cysteine residues) and thus it is not clear what drives metal transfer from one protein to the other. Cu transfer is a two-step reaction involving a metal-dependent ternary complex in which the metal is coordinated by cysteines from both proteins (i.e., Atox1-Cu-WD4). We employ size exclusion chromatography to estimate individual equilibrium constants for the two steps. This information together with calorimetric titration data are used to reveal enthalpic and entropic contributions of each step in the transfer process. Upon combining the equilibrium constants for both steps, a metal exchange factor (from Atox1 to WD4) of 10 is calculated, governed by a negative net enthalpy change of ∼10 kJ/mol. Thus, small variations in interaction energies, not always obvious upon comparing protein structures alone, may fuel vectorial metal transfer.
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spelling pubmed-33448372012-05-09 In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein Niemiec, Moritz S. Weise, Christoph F. Wittung-Stafshede, Pernilla PLoS One Research Article Transient protein-protein and protein-ligand interactions are fundamental components of biological activity. To understand biological activity, not only the structures of the involved proteins are important but also the energetics of the individual steps of a reaction. Here we use in vitro biophysical methods to deduce thermodynamic parameters of copper (Cu) transfer from the human copper chaperone Atox1 to the fourth metal-binding domain of the Wilson disease protein (WD4). Atox1 and WD4 have the same fold (ferredoxin-like fold) and Cu-binding site (two surface exposed cysteine residues) and thus it is not clear what drives metal transfer from one protein to the other. Cu transfer is a two-step reaction involving a metal-dependent ternary complex in which the metal is coordinated by cysteines from both proteins (i.e., Atox1-Cu-WD4). We employ size exclusion chromatography to estimate individual equilibrium constants for the two steps. This information together with calorimetric titration data are used to reveal enthalpic and entropic contributions of each step in the transfer process. Upon combining the equilibrium constants for both steps, a metal exchange factor (from Atox1 to WD4) of 10 is calculated, governed by a negative net enthalpy change of ∼10 kJ/mol. Thus, small variations in interaction energies, not always obvious upon comparing protein structures alone, may fuel vectorial metal transfer. Public Library of Science 2012-05-04 /pmc/articles/PMC3344837/ /pubmed/22574136 http://dx.doi.org/10.1371/journal.pone.0036102 Text en Niemiec et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Niemiec, Moritz S.
Weise, Christoph F.
Wittung-Stafshede, Pernilla
In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein
title In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein
title_full In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein
title_fullStr In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein
title_full_unstemmed In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein
title_short In Vitro Thermodynamic Dissection of Human Copper Transfer from Chaperone to Target Protein
title_sort in vitro thermodynamic dissection of human copper transfer from chaperone to target protein
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3344837/
https://www.ncbi.nlm.nih.gov/pubmed/22574136
http://dx.doi.org/10.1371/journal.pone.0036102
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