A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions
Chemokine receptors are extensively involved in a broad range of physiological and pathological processes, making them attractive drug targets. However, despite considerable efforts, there are very few approved drugs targeting this class of seven transmembrane domain receptors to date. In recent yea...
| Главные авторы: | , |
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| Формат: | Статья |
| Язык: | English |
| Опубликовано: |
MDPI AG
2022-04-01
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| Серии: | Cells |
| Предметы: | |
| Online-ссылка: | https://www.mdpi.com/2073-4409/11/8/1317 |
| _version_ | 1827621306353319936 |
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| author | Stefanie Alexandra Eberle Martin Gustavsson |
| author_facet | Stefanie Alexandra Eberle Martin Gustavsson |
| author_sort | Stefanie Alexandra Eberle |
| collection | DOAJ |
| description | Chemokine receptors are extensively involved in a broad range of physiological and pathological processes, making them attractive drug targets. However, despite considerable efforts, there are very few approved drugs targeting this class of seven transmembrane domain receptors to date. In recent years, the importance of including binding kinetics in drug discovery campaigns was emphasized. Therefore, kinetic insight into chemokine–chemokine receptor interactions could help to address this issue. Moreover, it could additionally deepen our understanding of the selectivity and promiscuity of the chemokine–chemokine receptor network. Here, we describe the application, optimization and validation of a homogenous Scintillation Proximity Assay (SPA) for real-time kinetic profiling of chemokine–chemokine receptor interactions on the example of ACKR3 and CXCL12. The principle of the SPA is the detection of radioligand binding to receptors reconstituted into nanodiscs by scintillation light. No receptor modifications are required. The nanodiscs provide a native-like environment for receptors and allow for full control over bilayer composition and size. The continuous assay format enables the monitoring of binding reactions in real-time, and directly accounts for non-specific binding and potential artefacts. Minor adaptations additionally facilitate the determination of equilibrium binding metrics, making the assay a versatile tool for the study of receptor–ligand interactions. |
| first_indexed | 2024-03-09T11:01:45Z |
| format | Article |
| id | doaj.art-fa95eedc56fa40b890db0199b901b475 |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-09T11:01:45Z |
| publishDate | 2022-04-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-fa95eedc56fa40b890db0199b901b4752023-12-01T01:14:15ZengMDPI AGCells2073-44092022-04-01118131710.3390/cells11081317A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor InteractionsStefanie Alexandra Eberle0Martin Gustavsson1Department of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3B, 2200 Copenhagen, DenmarkDepartment of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3B, 2200 Copenhagen, DenmarkChemokine receptors are extensively involved in a broad range of physiological and pathological processes, making them attractive drug targets. However, despite considerable efforts, there are very few approved drugs targeting this class of seven transmembrane domain receptors to date. In recent years, the importance of including binding kinetics in drug discovery campaigns was emphasized. Therefore, kinetic insight into chemokine–chemokine receptor interactions could help to address this issue. Moreover, it could additionally deepen our understanding of the selectivity and promiscuity of the chemokine–chemokine receptor network. Here, we describe the application, optimization and validation of a homogenous Scintillation Proximity Assay (SPA) for real-time kinetic profiling of chemokine–chemokine receptor interactions on the example of ACKR3 and CXCL12. The principle of the SPA is the detection of radioligand binding to receptors reconstituted into nanodiscs by scintillation light. No receptor modifications are required. The nanodiscs provide a native-like environment for receptors and allow for full control over bilayer composition and size. The continuous assay format enables the monitoring of binding reactions in real-time, and directly accounts for non-specific binding and potential artefacts. Minor adaptations additionally facilitate the determination of equilibrium binding metrics, making the assay a versatile tool for the study of receptor–ligand interactions.https://www.mdpi.com/2073-4409/11/8/13177TM receptorACKR3CXCL12SDF-1chemokinechemokine receptor |
| spellingShingle | Stefanie Alexandra Eberle Martin Gustavsson A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions Cells 7TM receptor ACKR3 CXCL12 SDF-1 chemokine chemokine receptor |
| title | A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions |
| title_full | A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions |
| title_fullStr | A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions |
| title_full_unstemmed | A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions |
| title_short | A Scintillation Proximity Assay for Real-Time Kinetic Analysis of Chemokine–Chemokine Receptor Interactions |
| title_sort | scintillation proximity assay for real time kinetic analysis of chemokine chemokine receptor interactions |
| topic | 7TM receptor ACKR3 CXCL12 SDF-1 chemokine chemokine receptor |
| url | https://www.mdpi.com/2073-4409/11/8/1317 |
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