Schaeferling, Michael and Wolfbeis, Otto S. (2007) Europium tetracycline as a luminescent probe for nucleoside phosphates and its application to the determination of kinase activity. CHEMISTRY-A EUROPEAN JOURNAL, 13 (15). pp. 4342-4349. ISSN 0947-6539,
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The determination of enzyme activities and the screening of enzyme regulators is a major task in clinical chemistry and drug development. A broad variety of enzymatic reactions is associated with the consumption of adenosine triphosphate (ATP), including, in particular, phosphorylation reactions catalyzed by kinases, formation of adenosine cyclic monophosphate (cAMP) by adenylate cyclases, and ATP decomposition by ATPase. We have studied the effect of a series of adenosine (ATP, ADP, AMP, cAMP) and guanosine (GTP, GDP) phosphoric esters, and of pyrophosphate (PP) on the fluorescence emission of the europium tetracycline (EuTC) complex. We found that these compounds have strongly different quenching effects on the luminescence emission of EuTC. ne triphosphates ATP and GTP behave as strong quenchers in reducing the fluorescence intensity of EuTC to 25% of its initial value by formation of a ternary 1:1:1 complex. All other phosphate esters showed a weak quenching effect only. The applicability of this fluorescent probe to the determination of the activity of phosphorylation enzymes is demonstrated by means of creatine kinase as a model for non-membrane-bound kinases. In contrast to other methods, this approach does not require the use of radioactively labeled ATP substrates, additional enzymes, or of rather complex immunoassays.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | TIME-RESOLVED FLUORESCENCE; NEUTRAL AQUEOUS-SOLUTION; HYDROGEN-PEROXIDE; ADENYLYL-CYCLASE; SPECTROFLUOROMETRIC DETERMINATION; MOLECULAR RECOGNITION; BIOANALYTICAL ASSAYS; ATP; COMPLEX; PYROPHOSPHATE; ATP; fluorescent probes; kinases; lanthanides; phosphorylation |
| Subjects: | 500 Science > 540 Chemistry & allied sciences |
| Divisions: | Chemistry and Pharmacy > Institut für Analytische Chemie, Chemo- und Biosensorik > Chemo- und Biosensorik (Prof. Antje J. Bäumner, formerly Prof. Wolfbeis) |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 11 Jan 2021 10:45 |
| Last Modified: | 11 Jan 2021 10:45 |
| URI: | https://pred.uni-regensburg.de/id/eprint/33364 |
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