title: Streamlined discovery of cross-linked chromatin complexes and associated histone modifications by mass spectrometry.
abstract: Posttranslational modifications (PTMs) are key contributors to chromatin function. The ability to comprehensively link specific histone PTMs with specific chromatin factors would be an important advance in understanding the functions and genomic targeting mechanisms of those factors. We recently introduced a cross-linked affinity technique, BioTAP-XL, to identify chromatin-bound protein interactions that can be difficult to capture with native affinity techniques. However, BioTAP-XL was not strictly compatible with similarly comprehensive analyses of associated histone PTMs. Here we advance BioTAP-XL by demonstrating the ability to quantify histone PTMs linked to specific chromatin factors in parallel with the ability to identify nonhistone binding partners. Furthermore we demonstrate that the initially published quantity of starting material can be scaled down orders of magnitude without loss in proteomic sensitivity. We also integrate hydrophilic interaction chromatography to mitigate detergent carryover and improve liquid chromatography-mass spectrometric performance. In summary, we greatly extend the practicality of BioTAP-XL to enable comprehensive identification of protein complexes and their local chromatin environment.
authors.affiliation: Division of Genetics, Brigham and Women's Hospital, Boston, MA 02115; Department of Genetics, Harvard Medical School, Boston, MA 02115;
authors.forename: Barry M
authors.initials: BM
authors.lastname: Zee
authors.affiliation: Division of Genetics, Brigham and Women's Hospital, Boston, MA 02115; Department of Genetics, Harvard Medical School, Boston, MA 02115;
authors.forename: Artyom A
authors.initials: AA
authors.lastname: Alekseyenko
authors.affiliation: Division of Genetics, Brigham and Women's Hospital, Boston, MA 02115; Department of Genetics, Harvard Medical School, Boston, MA 02115; Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138.
authors.forename: Kyle A
authors.initials: KA
authors.lastname: McElroy
authors.affiliation: Division of Genetics, Brigham and Women's Hospital, Boston, MA 02115; Department of Genetics, Harvard Medical School, Boston, MA 02115; mkuroda@genetics.med.harvard.edu.
authors.forename: Mitzi I
authors.initials: MI
authors.lastname: Kuroda
mesh_terms: D000818:Animals; D002843:Chromatin / Q000737:chemistry*; D002853:Chromatography, Liquid; D004330:Drosophila; D006657:Histones / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D040901:Proteomics
chemical_list: D002843:Chromatin; D006657:Histones
keywords: BioTAP-XL; affinity pulldown; chromatin complexes; histone modifications; liquid chromatography-mass spectrometry
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