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Title: Single-molecule kinetics and footprinting of DNA bis-intercalation: the paradigmatic case of Thiocoraline
Author: Camuñas Soler, Joan
Mañosas Castejón, María
Frutos, Silvia
Albericio Palomera, Fernando
Ritort Farran, Fèlix
Tulla-Puche, Judit
Keywords: Lligands (Bioquímica)
Ligands (Biochemistry)
Amino acids
Issue Date: 17-Feb-2015
Publisher: Oxford University Press
Abstract: DNA bis-intercalators are widely used in molecular biology with applications ranging from DNA imaging to anticancer pharmacology. Two fundamental aspects of these ligands are the lifetime of the bisintercalated complexes and their sequence selectivity. Here, we perform single-molecule optical tweezers experiments with the peptide Thiocoraline showing, for the first time, that bis-intercalation is driven by a very slow off-rate that steeply decreases with applied force. This feature reveals the existence of a long-lived (minutes) mono-intercalated intermediate that contributes to the extremely long lifetime of the complex (hours). We further exploit this particularly slow kinetics to determine the thermodynamics of binding and persistence length of bis-intercalated DNA for a given fraction of bound ligand, a measurement inaccessible in previous studies of faster intercalating agents. We also develop a novel singlemolecule footprinting technique based on DNA unzipping and determine the preferred binding sites of Thiocoraline with one base-pair resolution. This fast and radiolabelling-free footprinting technique provides direct access to the binding sites of small ligands to nucleic acids without the need of cleavage agents. Overall, our results provide new insights into the binding pathway of bis-intercalators and the reported selectivity might be of relevance for this and other anticancer drugs interfering with DNA replication and transcription in carcinogenic cell lines.
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It is part of: Nucleic Acids Research, 2015, vol. 43, num. 5, p. 2767-2779
ISSN: 0305-1048
Appears in Collections:Publicacions de projectes de recerca finançats per la UE
Articles publicats en revistes (Física de la Matèria Condensada)
Articles publicats en revistes (Química Inorgànica i Orgànica)

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