Forced intercalation as a tool in gene diagnostics and in studying DNA-protein interactions

Kohler, O., Jarikote, D. V., Singh, I. , Parmar, V. S., Weinhold, E. and Seitz, O. (2005) Forced intercalation as a tool in gene diagnostics and in studying DNA-protein interactions. Pure and Applied Chemistry, 77 (1). pp. 327-338. ISSN 0033-4545

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Aromatic and heteroaromatic groups that are forced to intercalate at specific positions in DNA are versatile probes of DNA-DNA and DNA-protein recognition. Fluorescent nucleobases are of value since they are able to report on localized alterations of DNA duplex structure. However, the fluorescence of the vast majority of base surrogates becomes quenched upon intercalation in DNA. Peptide nucleic acid (PNA)-based probes are presented in which the intercalator dye thiazole orange (TO) serves as a fluorescent base surrogate. In these probes, fluorescence increases (5-60-fold) upon hybridization. PNA-bearing TO as fluorescent base surrogate could hence prove useful in real-time polymerase chain reaction (PCR) applications and in live cell analysis. Forced intercalation of aromatic polycycles can help to explore the binding mechanism of DNA-modifying enzymes. We discuss studies of DNA-methyltransferases (MTases) which commence methylation of nucleobases in DNA by flipping the target nucleotide completely out of the helix. A method for probing the baseflipping mechanism is suggested. It draws upon the observation that large hydrophobic base surrogates in the face of the swung-out base can enhance the DNA-enzyme binding affinity possibly by disrupting target base-stacking and stabilizing the apparent abasic site.

Subjects:B Subjects allied to Medicine > B200 Pharmacology, Toxicology and Pharmacy
Divisions:College of Science > School of Pharmacy
ID Code:11304
Deposited On:24 Jul 2013 15:40

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