GFP-like fluorophores as DNA labels for studying DNA-protein interactions
Language English Country United States Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
PubMed
22935023
DOI
10.1021/jo301684b
Knihovny.cz E-resources
- MeSH
- Benzyl Compounds chemistry MeSH
- DNA-Binding Proteins chemistry metabolism MeSH
- DNA-Directed DNA Polymerase chemistry metabolism MeSH
- DNA chemistry metabolism MeSH
- Fluorescent Dyes chemistry MeSH
- Imidazolines MeSH
- Imidazoles chemistry MeSH
- Molecular Sequence Data MeSH
- Nucleotides chemistry metabolism MeSH
- Base Pairing MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- 3,5-bis(methoxy)-4-hydroxy-benzylideneimidazolinone MeSH Browser
- 3,5-difluoro-4-hydroxybenzylidene imidazolinone MeSH Browser
- Benzyl Compounds MeSH
- DNA-Binding Proteins MeSH
- DNA-Directed DNA Polymerase MeSH
- DNA MeSH
- Fluorescent Dyes MeSH
- Imidazolines MeSH
- Imidazoles MeSH
- Nucleotides MeSH
- Tli polymerase MeSH Browser
GFP-like 3,5-difluoro-4-hydroxybenzylideneimidazolinone (FBI) and 3,5-bis(methoxy)-4-hydroxy-benzylideneimidazolinone (MBI) labels were attached to dCTP through a propargyl linker, and the resulting labeled nucleotides (dC(MBI)TP and dC(FBI)TP) were used for a facile enzymatic synthesis of oligonucleotide or DNA probes by polymerase-catalyzed primer extension. The MBI/FBI-labeled DNA probes exerted low fluorescence that was increased 2-3.2 times upon binding of a protein. The concept was demonstrated on sequence-specific binding of p53 to dsDNA and on nonspecific binding of single strand binding protein to an oligonucleotide. The FBI label was also used for a time-resolved experiment monitoring a single-nucleotide incorporation followed by primer extension by Vent(exo-) polymerase.
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