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Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein

We present a new methodology for site-specific sensing of peptide–oligonucleotide (ODN) interactions using a solvatochromic fluorescent label based on 3-hydroxychromone (3HC). This label was covalently attached to the N-terminus of a peptide corresponding to the zinc finger domain of the HIV-1 nucle...

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Autores principales: Shvadchak, Volodymyr V., Klymchenko, Andrey S., de Rocquigny, Hugues, Mély, Yves
Formato: Texto
Lenguaje:English
Publicado: Oxford University Press 2009
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2647317/
https://www.ncbi.nlm.nih.gov/pubmed/19151084
http://dx.doi.org/10.1093/nar/gkn1083
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author Shvadchak, Volodymyr V.
Klymchenko, Andrey S.
de Rocquigny, Hugues
Mély, Yves
author_facet Shvadchak, Volodymyr V.
Klymchenko, Andrey S.
de Rocquigny, Hugues
Mély, Yves
author_sort Shvadchak, Volodymyr V.
collection PubMed
description We present a new methodology for site-specific sensing of peptide–oligonucleotide (ODN) interactions using a solvatochromic fluorescent label based on 3-hydroxychromone (3HC). This label was covalently attached to the N-terminus of a peptide corresponding to the zinc finger domain of the HIV-1 nucleocapsid protein (NC). On interaction with target ODNs, the labeled peptide shows strong changes in the ratio of its two emission bands, indicating an enhanced screening of the 3HC fluorophore from the bulk water by the ODN bases. Remarkably, this two-color response depends on the ODN sequence and correlates with the 3D structure of the corresponding complexes, suggesting that the 3HC label monitors the peptide–ODN interactions site-specifically. By measuring the two-color ratio, we were also able to determine the peptide–ODN-binding parameters and distinguish multiple binding sites in ODNs, which is rather difficult using other fluorescence methods. Moreover, this method was found to be more sensitive than the commonly used steady-state fluorescence anisotropy, especially in the case of small ODNs. The described methodology could become a new universal tool for investigating peptide–ODN interactions.
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spelling pubmed-26473172009-03-04 Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein Shvadchak, Volodymyr V. Klymchenko, Andrey S. de Rocquigny, Hugues Mély, Yves Nucleic Acids Res Methods Online We present a new methodology for site-specific sensing of peptide–oligonucleotide (ODN) interactions using a solvatochromic fluorescent label based on 3-hydroxychromone (3HC). This label was covalently attached to the N-terminus of a peptide corresponding to the zinc finger domain of the HIV-1 nucleocapsid protein (NC). On interaction with target ODNs, the labeled peptide shows strong changes in the ratio of its two emission bands, indicating an enhanced screening of the 3HC fluorophore from the bulk water by the ODN bases. Remarkably, this two-color response depends on the ODN sequence and correlates with the 3D structure of the corresponding complexes, suggesting that the 3HC label monitors the peptide–ODN interactions site-specifically. By measuring the two-color ratio, we were also able to determine the peptide–ODN-binding parameters and distinguish multiple binding sites in ODNs, which is rather difficult using other fluorescence methods. Moreover, this method was found to be more sensitive than the commonly used steady-state fluorescence anisotropy, especially in the case of small ODNs. The described methodology could become a new universal tool for investigating peptide–ODN interactions. Oxford University Press 2009-02 2009-01-16 /pmc/articles/PMC2647317/ /pubmed/19151084 http://dx.doi.org/10.1093/nar/gkn1083 Text en © 2009 The Author(s) http://creativecommons.org/licenses/by-nc/2.0/uk/ This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Methods Online
Shvadchak, Volodymyr V.
Klymchenko, Andrey S.
de Rocquigny, Hugues
Mély, Yves
Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein
title Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein
title_full Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein
title_fullStr Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein
title_full_unstemmed Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein
title_short Sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the HIV-1 nucleocapsid protein
title_sort sensing peptide–oligonucleotide interactions by a two-color fluorescence label: application to the hiv-1 nucleocapsid protein
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2647317/
https://www.ncbi.nlm.nih.gov/pubmed/19151084
http://dx.doi.org/10.1093/nar/gkn1083
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