Department of Molecular Biology, Radboud Institute for Molecular Life Sciences, Radboud University, Nijmegen, 6500 HB, The Netherlands.
Oncode Institute, Radboud University, Nijmegen, 6500 HB, The Netherlands.
Nat Commun. 2018 Apr 25;9(1):1653. doi: 10.1038/s41467-018-04084-0.
Interaction proteomics studies have provided fundamental insights into multimeric biomolecular assemblies and cell-scale molecular networks. Significant recent developments in mass spectrometry-based interaction proteomics have been fueled by rapid advances in label-free, isotopic, and isobaric quantitation workflows. Here, we report a quantitative protein-DNA and protein-nucleosome binding assay that uses affinity purifications from nuclear extracts coupled with isobaric chemical labeling and mass spectrometry to quantify apparent binding affinities proteome-wide. We use this assay with a variety of DNA and nucleosome baits to quantify apparent binding affinities of monomeric and multimeric transcription factors and chromatin remodeling complexes.
相互作用蛋白质组学研究为多聚生物分子组装体和细胞尺度分子网络提供了基本的见解。基于质谱的相互作用蛋白质组学的重大进展得益于无标记、同位素和等压定量工作流程的快速发展。在这里，我们报告了一种定量的蛋白质-DNA 和蛋白质核小体结合测定法，该方法使用核提取物的亲和纯化，结合等压化学标记和质谱，在全蛋白质组范围内定量表观结合亲和力。我们使用该测定法和各种 DNA 和核小体诱饵，来定量单体和多聚体转录因子和染色质重塑复合物的表观结合亲和力。
