Archipowa, Nataliya and Wittmann, Lukas and Koeckenberger, Johannes and Ertl, Fabian J. and Gleixner, Jakob and Keller, Max and Heinrich, Markus R. and Kutta, Roger Jan (2023) Characterization of Fluorescent Dyes Frequently Used for Bioimaging: Photophysics and Photocatalytical Reactions with Proteins. JOURNAL OF PHYSICAL CHEMISTRY B, 127 (44). pp. 9532-9542. ISSN 1520-6106, 1520-5207
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Derivatives of the rhodamine-based dye 5-TAMRA (5-carboxy-tetramethylrhodamine) and the indocarbocyanine-type Cy3B (cyclized derivative of the cyanine dye Cy3), both representing important fluorophores frequently used for the labeling of biomolecules (proteins, nucleic acids) and bioactive compounds, such as receptor ligands, were photophysically investigated in aqueous solution, i.e., in neat phosphate-buffered saline (PBS) and in PBS supplemented with 1 wt % bovine serum albumin (BSA). The dyes exhibit comparable absorption (lambda(abs,max): 550-569 nm) and emission wavelengths (lambda(em,max): 580-582 nm), and similar S-1 lifetimes (2.27-2.75 ns), and their excited state deactivation proceeds mainly via the lowest excited singlet state (triplet quantum yield ca. 1%). However, the probes show marked differences with respect to their fluorescence quantum yield and photostability. While 5-TAMRA shows a lower quantum yield (37-39%) than the Cy3B derivative (ca. 57%), its photostability is considerably higher compared to Cy3B. Generally, the impact of the protein on the photophysics is low. However, on prolonged illumination, both fluorescent dyes undergo a photocatalytic reaction with tryptophan residues of BSA mediated by sensitized singlet oxygen resulting in a tryptophan photoproduct with an absorption maximum around 330 nm. The overall results of this work will assist in choosing the right dye for the labeling of bioactive compounds, and the study demonstrates that experiments performed with 5-TAMRA or Cy3B-labeled compounds in a biological environment may be influenced by photochemical modification of experimentally relevant proteins at aromatic amino acid residues.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | COUPLED RECEPTORS; TRYPTOPHAN; PHARMACOLOGY; PERFORMANCE; DEGRADATION; CONVENIENT; LIGANDS; |
| Subjects: | 500 Science > 540 Chemistry & allied sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Reinhard Sterner Chemistry and Pharmacy > Institute of Pharmacy > Pharmaceutical/Medicinal Chemistry II (Prof. Buschauer) Chemistry and Pharmacy > Institut für Physikalische und Theoretische Chemie Chemistry and Pharmacy > Institut für Physikalische und Theoretische Chemie > Chair of Physical Chemistry I > Prof. Dr. Patrick Nürnberger |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 09 Mar 2024 14:46 |
| Last Modified: | 09 Mar 2024 14:47 |
| URI: | https://pred.uni-regensburg.de/id/eprint/59414 |
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