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The Effects of Sodium Ions on Ligand Binding and Conformational States of G Protein-Coupled Receptors—Insights from Mass Spectrometry
[Image: see text] The use of mass spectrometry to investigate proteins is now well established and provides invaluable information for both soluble and membrane protein assemblies. Maintaining transient noncovalent interactions under physiological conditions, however, remains challenging. Here, usin...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Chemical
Society
2021
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7995251/ https://www.ncbi.nlm.nih.gov/pubmed/33711230 http://dx.doi.org/10.1021/jacs.0c11837 |
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author | Agasid, Mark T. Sørensen, Lars Urner, Leonhard H. Yan, Jun Robinson, Carol V. |
author_facet | Agasid, Mark T. Sørensen, Lars Urner, Leonhard H. Yan, Jun Robinson, Carol V. |
author_sort | Agasid, Mark T. |
collection | PubMed |
description | [Image: see text] The use of mass spectrometry to investigate proteins is now well established and provides invaluable information for both soluble and membrane protein assemblies. Maintaining transient noncovalent interactions under physiological conditions, however, remains challenging. Here, using nanoscale electrospray ionization emitters, we establish conditions that enable mass spectrometry of two G protein-coupled receptors (GPCR) from buffers containing high concentrations of sodium ions. For the Class A GPCR, the adenosine 2A receptor, we observe ligand-induced changes to sodium binding of the receptor at the level of individual sodium ions. We find that antagonists promote sodium binding while agonists attenuate sodium binding. These findings are in line with high-resolution X-ray crystallography wherein only inactive conformations retain sodium ions in allosteric binding pockets. For the glucagon receptor (a Class B GPCR) we observed enhanced ligand binding in electrospray buffers containing high concentrations of sodium, as opposed to ammonium acetate buffers. A combination of native and -omics mass spectrometry revealed the presence of a lipophilic negative allosteric modulator. These experiments highlight the advantages of implementing native mass spectrometry, from electrospray buffers containing high concentrations of physiologically relevant salts, to inform on allosteric ions or ligands with the potential to define their roles on GPCR function. |
format | Online Article Text |
id | pubmed-7995251 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | American Chemical
Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-79952512021-03-29 The Effects of Sodium Ions on Ligand Binding and Conformational States of G Protein-Coupled Receptors—Insights from Mass Spectrometry Agasid, Mark T. Sørensen, Lars Urner, Leonhard H. Yan, Jun Robinson, Carol V. J Am Chem Soc [Image: see text] The use of mass spectrometry to investigate proteins is now well established and provides invaluable information for both soluble and membrane protein assemblies. Maintaining transient noncovalent interactions under physiological conditions, however, remains challenging. Here, using nanoscale electrospray ionization emitters, we establish conditions that enable mass spectrometry of two G protein-coupled receptors (GPCR) from buffers containing high concentrations of sodium ions. For the Class A GPCR, the adenosine 2A receptor, we observe ligand-induced changes to sodium binding of the receptor at the level of individual sodium ions. We find that antagonists promote sodium binding while agonists attenuate sodium binding. These findings are in line with high-resolution X-ray crystallography wherein only inactive conformations retain sodium ions in allosteric binding pockets. For the glucagon receptor (a Class B GPCR) we observed enhanced ligand binding in electrospray buffers containing high concentrations of sodium, as opposed to ammonium acetate buffers. A combination of native and -omics mass spectrometry revealed the presence of a lipophilic negative allosteric modulator. These experiments highlight the advantages of implementing native mass spectrometry, from electrospray buffers containing high concentrations of physiologically relevant salts, to inform on allosteric ions or ligands with the potential to define their roles on GPCR function. American Chemical Society 2021-03-12 2021-03-24 /pmc/articles/PMC7995251/ /pubmed/33711230 http://dx.doi.org/10.1021/jacs.0c11837 Text en © 2021 The Authors. Published byAmerican Chemical Society Permits the broadest form of re-use including for commercial purposes, provided that author attribution and integrity are maintained (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Agasid, Mark T. Sørensen, Lars Urner, Leonhard H. Yan, Jun Robinson, Carol V. The Effects of Sodium Ions on Ligand Binding and Conformational States of G Protein-Coupled Receptors—Insights from Mass Spectrometry |
title | The
Effects of Sodium Ions on Ligand Binding and Conformational
States of G Protein-Coupled Receptors—Insights from Mass Spectrometry |
title_full | The
Effects of Sodium Ions on Ligand Binding and Conformational
States of G Protein-Coupled Receptors—Insights from Mass Spectrometry |
title_fullStr | The
Effects of Sodium Ions on Ligand Binding and Conformational
States of G Protein-Coupled Receptors—Insights from Mass Spectrometry |
title_full_unstemmed | The
Effects of Sodium Ions on Ligand Binding and Conformational
States of G Protein-Coupled Receptors—Insights from Mass Spectrometry |
title_short | The
Effects of Sodium Ions on Ligand Binding and Conformational
States of G Protein-Coupled Receptors—Insights from Mass Spectrometry |
title_sort | the
effects of sodium ions on ligand binding and conformational
states of g protein-coupled receptors—insights from mass spectrometry |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7995251/ https://www.ncbi.nlm.nih.gov/pubmed/33711230 http://dx.doi.org/10.1021/jacs.0c11837 |
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