PUBLICATION
A novel molecular rotor facilitates detection of p53-DNA interactions using the Fluorescent Intercalator Displacement Assay
- Authors
- Goh, W.L., Lee, M.Y., Lim, T.X., Chua, J.S., Brenner, S., Ghadessy, F.J., Teo, Y.N.
- ID
- ZDB-PUB-180830-3
- Date
- 2018
- Source
- Scientific Reports 8: 12946 (Journal)
- Registered Authors
- Keywords
- none
- MeSH Terms
-
- DNA/chemistry*
- DNA/metabolism
- Fluorescent Dyes/chemistry*
- Humans
- Intercalating Agents/chemistry*
- Spectrometry, Fluorescence
- Tumor Suppressor Protein p53/chemistry*
- Tumor Suppressor Protein p53/metabolism
- PubMed
- 30154420 Full text @ Sci. Rep.
Citation
Goh, W.L., Lee, M.Y., Lim, T.X., Chua, J.S., Brenner, S., Ghadessy, F.J., Teo, Y.N. (2018) A novel molecular rotor facilitates detection of p53-DNA interactions using the Fluorescent Intercalator Displacement Assay. Scientific Reports. 8:12946.
Abstract
We have investigated the use of fluorescent molecular rotors as probes for detection of p53 binding to DNA. These are a class of fluorophores that undergo twisted intramolecular charge transfer (TICT). They are non-fluorescent in a freely rotating conformation and experience a fluorescence increase when restricted in the planar conformation. We hypothesized that intercalation of a molecular rotor between DNA base pairs would result in a fluorescence turn-on signal. Upon displacement by a DNA binding protein, measurable loss of signal would facilitate use of the molecular rotor in the fluorescent intercalator displacement (FID) assay. A panel of probes was interrogated using the well-established p53 model system across various DNA response elements. A novel, readily synthesizable molecular rotor incorporating an acridine orange DNA intercalating group (AO-R) outperformed other conventional dyes in the FID assay. It enabled relative measurement of p53 sequence-specific DNA interactions and study of the dominant-negative effects of cancer-associated p53 mutants. In a further application, AO-R also proved useful for staining apoptotic cells in live zebrafish embryos.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping