Proteomics

Dataset Information

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H4K20me2 SILAC Nucleosome Affinity Purification


ABSTRACT: Goal of this project was the identification of chromatin interacting proteins whose binding is differentially regulated by di-methylation of lysine 20 on histone H4 (H4K20me2). To achieve this unodified and H4K20me2-modified histone H4 were generated by native chemical ligation and assembled into recombinant di-nucleosomes. The di-nucleosomes were immobilized on streptavidin-coated beads via the biotinylated di-nucleosomal DNA and used for nucleosome affinity purifications to identify proteins regulated by H4K20me2 from SILAC-labelled HeLaS3 nuclear extracts (Arg10 and Lys8). SILAC affinity purifications were carried out in "forward" (heavy extract on modified nucleosome and light extract on unmodified nucleosome) and "reverse" (light extract on modified nucleosome and heavy extract on unmodified nucleosome) label-swap experiments and protein abundances were quantified by MaxQuant.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Hela Cell

SUBMITTER: Saulius Lukauskas  

LAB HEAD: Till Bartke

PROVIDER: PXD009281 | Pride | 2019-02-27

REPOSITORIES: Pride

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Publications


Genotoxic DNA double-strand breaks (DSBs) can be repaired by error-free homologous recombination (HR) or mutagenic non-homologous end-joining<sup>1</sup>. HR supresses tumorigenesis<sup>1</sup>, but is restricted to the S and G2 phases of the cell cycle when a sister chromatid is present<sup>2</sup>. Breast cancer type 1 susceptibility protein (BRCA1) promotes HR by antagonizing the anti-resection factor TP53-binding protein 1(53BP1) (refs. <sup>2-5</sup>), but it remains unknown how BRCA1 funct  ...[more]

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