Analysis of protein-DNA interactions in chromatin by UV induced cross-linking and mass spectrometry

Alexandra Stützer, Luisa M. Welp, Monika Raabe, Timo Sachsenberg, Christin Kappert, Alexander Wulf, Andy M. Lau, Stefan-Sebastian David, Aleksandar Chernev, Katharina Kramer, Argyris Politis, Oliver Kohlbacher, Wolfgang Fischle and Henning Urlaub ()
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Alexandra Stützer: Max Planck Institute for Biophysical Chemistry
Luisa M. Welp: Max Planck Institute for Biophysical Chemistry
Monika Raabe: Max Planck Institute for Biophysical Chemistry
Timo Sachsenberg: University of Tübingen
Christin Kappert: Max Planck Institute for Biophysical Chemistry
Alexander Wulf: Max Planck Institute for Biophysical Chemistry
Andy M. Lau: King’s College London
Stefan-Sebastian David: Max Planck Institute for Biophysical Chemistry
Aleksandar Chernev: Max Planck Institute for Biophysical Chemistry
Katharina Kramer: CSL Behring GmbH
Argyris Politis: King’s College London
Oliver Kohlbacher: University of Tübingen
Wolfgang Fischle: Max Planck Institute for Biophysical Chemistry
Henning Urlaub: Max Planck Institute for Biophysical Chemistry

Nature Communications, 2020, vol. 11, issue 1, 1-12

Abstract: Abstract Protein–DNA interactions are key to the functionality and stability of the genome. Identification and mapping of protein–DNA interaction interfaces and sites is crucial for understanding DNA-dependent processes. Here, we present a workflow that allows mass spectrometric (MS) identification of proteins in direct contact with DNA in reconstituted and native chromatin after cross-linking by ultraviolet (UV) light. Our approach enables the determination of contact interfaces at amino-acid level. With the example of chromatin-associated protein SCML2 we show that our technique allows differentiation of nucleosome-binding interfaces in distinct states. By UV cross-linking of isolated nuclei we determined the cross-linking sites of several factors including chromatin-modifying enzymes, demonstrating that our workflow is not restricted to reconstituted materials. As our approach can distinguish between protein–RNA and DNA interactions in one single experiment, we project that it will be possible to obtain insights into chromatin and its regulation in the future.

Date: 2020
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Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:11:y:2020:i:1:d:10.1038_s41467-020-19047-7

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DOI: 10.1038/s41467-020-19047-7

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