Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions
Analyses of protein interactions are fundamental for the investigation of molecular mechanisms responsible for cellular processes and diseases, as well as for drug discovery in the pharmaceutical industry. The present study details the development of a fluorescence polarization assay using melanoma...
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2009-10-01
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| Series: | BioTechniques |
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| Online Access: | https://www.future-science.com/doi/10.2144/000113223 |
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| author | Alexander Riechers Jennifer Schmidt Burkhard König Anja Katrin Bosserhoff |
| author_facet | Alexander Riechers Jennifer Schmidt Burkhard König Anja Katrin Bosserhoff |
| author_sort | Alexander Riechers |
| collection | DOAJ |
| description | Analyses of protein interactions are fundamental for the investigation of molecular mechanisms responsible for cellular processes and diseases, as well as for drug discovery in the pharmaceutical industry. The present study details the development of a fluorescence polarization assay using melanoma inhibitory activity (MIA) protein–binding compounds and studies of the binding properties of this protein. Since they are dependent on the the lifetime of the fluorescent label, currently available fluorescence polarization assays can only determine interactions with either high– or low–molecular weight interaction partners. Our new approach eliminates this limitation by immobilizing a known binding partner of MIA protein to a well plate and by labeling the target protein using luminescent transition metal labels such as Ru(bpy)3 for binding studies with both high– and low–molecular weight interaction partners. Due to the use of a functionalized surface, we termed our concept heterogeneous transition metal–based fluorescence polarization (HTFP) assay. The assay's independence from the molecular weight of potential binding partners should make the technique amenable to investigations on subjects as diverse as multimerization, interactions with pharmacophores, or binding affinity determination. |
| format | Article |
| id | doaj-art-9c2bd190ca4e41a6897aed11b55d837e |
| institution | OA Journals |
| issn | 0736-6205 1940-9818 |
| language | English |
| publishDate | 2009-10-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | BioTechniques |
| spelling | doaj-art-9c2bd190ca4e41a6897aed11b55d837e2025-08-20T02:25:51ZengTaylor & Francis GroupBioTechniques0736-62051940-98182009-10-0147483784410.2144/000113223Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactionsAlexander Riechers0Jennifer Schmidt1Burkhard König2Anja Katrin Bosserhoff31Institute for Organic Chemistry, University of Regensburg, Regensburg, Germany2Institute of Pathology, University of Regensburg, Regensburg, Germany1Institute for Organic Chemistry, University of Regensburg, Regensburg, Germany2Institute of Pathology, University of Regensburg, Regensburg, GermanyAnalyses of protein interactions are fundamental for the investigation of molecular mechanisms responsible for cellular processes and diseases, as well as for drug discovery in the pharmaceutical industry. The present study details the development of a fluorescence polarization assay using melanoma inhibitory activity (MIA) protein–binding compounds and studies of the binding properties of this protein. Since they are dependent on the the lifetime of the fluorescent label, currently available fluorescence polarization assays can only determine interactions with either high– or low–molecular weight interaction partners. Our new approach eliminates this limitation by immobilizing a known binding partner of MIA protein to a well plate and by labeling the target protein using luminescent transition metal labels such as Ru(bpy)3 for binding studies with both high– and low–molecular weight interaction partners. Due to the use of a functionalized surface, we termed our concept heterogeneous transition metal–based fluorescence polarization (HTFP) assay. The assay's independence from the molecular weight of potential binding partners should make the technique amenable to investigations on subjects as diverse as multimerization, interactions with pharmacophores, or binding affinity determination.https://www.future-science.com/doi/10.2144/000113223fluorescence polarizationprotein interactionsluminescent label |
| spellingShingle | Alexander Riechers Jennifer Schmidt Burkhard König Anja Katrin Bosserhoff Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions BioTechniques fluorescence polarization protein interactions luminescent label |
| title | Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions |
| title_full | Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions |
| title_fullStr | Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions |
| title_full_unstemmed | Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions |
| title_short | Heterogeneous transition metal–based fluorescence polarization (HTFP) assay for probing protein interactions |
| title_sort | heterogeneous transition metal based fluorescence polarization htfp assay for probing protein interactions |
| topic | fluorescence polarization protein interactions luminescent label |
| url | https://www.future-science.com/doi/10.2144/000113223 |
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