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Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method

Mass spectrometry is a powerful alternative to antibody-based methods for the analysis of histone post-translational modifications (marks). A key development in this approach was the deliberate propionylation of histones to improve sequence coverage across the lysine-rich and hydrophilic tails that...

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Autores principales: Maile, Tobias M., Izrael-Tomasevic, Anita, Cheung, Tommy, Guler, Gulfem D., Tindell, Charles, Masselot, Alexandre, Liang, Jun, Zhao, Feng, Trojer, Patrick, Classon, Marie, Arnott, David
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Biochemistry and Molecular Biology 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4390259/
https://www.ncbi.nlm.nih.gov/pubmed/25680960
http://dx.doi.org/10.1074/mcp.O114.046573
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author Maile, Tobias M.
Izrael-Tomasevic, Anita
Cheung, Tommy
Guler, Gulfem D.
Tindell, Charles
Masselot, Alexandre
Liang, Jun
Zhao, Feng
Trojer, Patrick
Classon, Marie
Arnott, David
author_facet Maile, Tobias M.
Izrael-Tomasevic, Anita
Cheung, Tommy
Guler, Gulfem D.
Tindell, Charles
Masselot, Alexandre
Liang, Jun
Zhao, Feng
Trojer, Patrick
Classon, Marie
Arnott, David
author_sort Maile, Tobias M.
collection PubMed
description Mass spectrometry is a powerful alternative to antibody-based methods for the analysis of histone post-translational modifications (marks). A key development in this approach was the deliberate propionylation of histones to improve sequence coverage across the lysine-rich and hydrophilic tails that bear most modifications. Several marks continue to be problematic however, particularly di- and tri-methylated lysine 4 of histone H3 which we found to be subject to substantial and selective losses during sample preparation and liquid chromatography-mass spectrometry. We developed a new method employing a “one-pot” hybrid chemical derivatization of histones, whereby an initial conversion of free lysines to their propionylated forms under mild aqueous conditions is followed by trypsin digestion and labeling of new peptide N termini with phenyl isocyanate. High resolution mass spectrometry was used to collect qualitative and quantitative data, and a novel web-based software application (Fishtones) was developed for viewing and quantifying histone marks in the resulting data sets. Recoveries of 53 methyl, acetyl, and phosphoryl marks on histone H3.1 were improved by an average of threefold overall, and over 50-fold for H3K4 di- and tri-methyl marks. The power of this workflow for epigenetic research and drug discovery was demonstrated by measuring quantitative changes in H3K4 trimethylation induced by small molecule inhibitors of lysine demethylases and siRNA knockdown of epigenetic modifiers ASH2L and WDR5.
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spelling pubmed-43902592015-04-17 Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method Maile, Tobias M. Izrael-Tomasevic, Anita Cheung, Tommy Guler, Gulfem D. Tindell, Charles Masselot, Alexandre Liang, Jun Zhao, Feng Trojer, Patrick Classon, Marie Arnott, David Mol Cell Proteomics Technological Innovation and Resources Mass spectrometry is a powerful alternative to antibody-based methods for the analysis of histone post-translational modifications (marks). A key development in this approach was the deliberate propionylation of histones to improve sequence coverage across the lysine-rich and hydrophilic tails that bear most modifications. Several marks continue to be problematic however, particularly di- and tri-methylated lysine 4 of histone H3 which we found to be subject to substantial and selective losses during sample preparation and liquid chromatography-mass spectrometry. We developed a new method employing a “one-pot” hybrid chemical derivatization of histones, whereby an initial conversion of free lysines to their propionylated forms under mild aqueous conditions is followed by trypsin digestion and labeling of new peptide N termini with phenyl isocyanate. High resolution mass spectrometry was used to collect qualitative and quantitative data, and a novel web-based software application (Fishtones) was developed for viewing and quantifying histone marks in the resulting data sets. Recoveries of 53 methyl, acetyl, and phosphoryl marks on histone H3.1 were improved by an average of threefold overall, and over 50-fold for H3K4 di- and tri-methyl marks. The power of this workflow for epigenetic research and drug discovery was demonstrated by measuring quantitative changes in H3K4 trimethylation induced by small molecule inhibitors of lysine demethylases and siRNA knockdown of epigenetic modifiers ASH2L and WDR5. The American Society for Biochemistry and Molecular Biology 2015-04 2015-02-13 /pmc/articles/PMC4390259/ /pubmed/25680960 http://dx.doi.org/10.1074/mcp.O114.046573 Text en © 2015 by The American Society for Biochemistry and Molecular Biology, Inc. Author's Choice—Final version full access.
spellingShingle Technological Innovation and Resources
Maile, Tobias M.
Izrael-Tomasevic, Anita
Cheung, Tommy
Guler, Gulfem D.
Tindell, Charles
Masselot, Alexandre
Liang, Jun
Zhao, Feng
Trojer, Patrick
Classon, Marie
Arnott, David
Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method
title Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method
title_full Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method
title_fullStr Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method
title_full_unstemmed Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method
title_short Mass Spectrometric Quantification of Histone Post-translational Modifications by a Hybrid Chemical Labeling Method
title_sort mass spectrometric quantification of histone post-translational modifications by a hybrid chemical labeling method
topic Technological Innovation and Resources
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4390259/
https://www.ncbi.nlm.nih.gov/pubmed/25680960
http://dx.doi.org/10.1074/mcp.O114.046573
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