| Literature DB >> 8202510 |
Abstract
A method is described for monitoring the enzyme-mediated unwinding of duplex DNA spectrophotometrically. The assay employs a partially duplex oligonucleotide substrate modified at the complementary end with coumarin and fluorescein moieties. When in close proximity the fluorescein quenches the fluorescence of coumarin. However, when the strands are separated by the action of a DNA helicase, the coumarin fluorescence increases greatly. Therefore, the progress of enzyme-mediated DNA unwinding can be measured in real time by fluorescence spectroscopy. This assay provides a simple method to screen for helicase inhibitors, which are of growing interest as potential anticancer agents. The application of this technique to kinetic analyses of the mechanism of action of DNA helicases is also discussed.Entities:
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Year: 1994 PMID： 8202510 PMCID： PMC44017 DOI： 10.1073/pnas.91.12.5471
Source DB: PubMed Journal: Proc Natl Acad Sci U S A ISSN： 0027-8424 Impact factor: 11.205