Specific protein-binding reactions monitored with ligand-ATP conjugates and firefly luciferase
| Autor: | Robert C. Boguslaski, Robert J. Carrico, James Edward Christner, Robert T. Buckler, Hartmut R. Schroeder, Kwok-Kam Yeung |
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| Rok vydání: | 1976 |
| Předmět: |
Specific protein
biology Chemistry Diptera Biophysics Cell Biology Ligands Ligand (biochemistry) Biochemistry Antigen-Antibody Reactions Kinetics Residue (chemistry) Adenosine Triphosphate Covalent bond Methods biology.protein Luciferase Antibody Luciferases Molecular Biology Incubation Protein Binding Conjugate |
| Zdroj: | Analytical Biochemistry. 76:95-110 |
| ISSN: | 0003-2697 |
| DOI: | 10.1016/0003-2697(76)90267-0 |
| Popis: | A new method was developed to monitor specific protein binding reactions with an ATP-labeled ligand and firefly luciferase. The ligand, 2,4-dinitrobenzene, was covalently coupled to four ATP derivatives and three of these conjugates were measured quantitatively at nanomolar levels with firefly luciferase. Incubation of the conjugates with antibody to the 2,4-dinitrophenyl residue diminished the peak light intensities produced in the bioluminescent assay, whereas incubation with immunoglobulin from a nonimmunized rabbit did not affect light production. Therefore, the antibody-bound ligand-ATP conjugates were inactive in the bioluminescent assay and levels of unbound conjugate could be measured in the presence of the bound form. The firefly luciferase was used to monitor competitive binding reactions between the antibody, the conjugates, and N(2,4-dinitrophenyl)-β-alanine. |
| Databáze: | OpenAIRE |
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