Year: 2026
Type: article
Abstract: Nonenzymatic covalent modification (NECM) of lysine residues can be physiologically consequential. A NECM is formed by the oxidized abasic site (C4-AP), which is produced by DNA-damaging agents. C4-AP reacts with the ε-amine of histone lysines in nucleosome core particles (NCPs) to form an electrophilic 5-methylene pyrrolone NECM (K MP ). K MP is also produced on histones in bleomycin-treated human cells. Here, we describe a molecule ( 1a ) that yields K MP by reacting directly with histones in NCPs. K MP forms on lysines of all four core histones in the order H3 > H2A/H2B > H4. Biotinylated K MP -containing NCPs prepared using 1a were incubated with HeLa nuclear lysates in the presence of glutathione. NCP–protein cross-links were observed by native PAGE. Protein–protein cross-links (PPCs) were enriched through intact NCP pull-down and identified via tryptic digests by LC-MS/MS. Model reactions demonstrate K MP is more electrophilic than N -acyllysine post-translational modifications (PTMs) but does not form PPCs indiscriminately within NCPs. Enriched proteins are functionally biased, with overrepresentation of DNA binding, histone binding, histone PTMs, and transcription regulation. Proteins enriched by K MP -containing NCPs produced by generating C4-AP on DNA were analyzed in parallel. Similar overrepresented functions were observed when C4-AP was introduced near the H3/H4 N-tails, whereas a distinct group of proteins was enriched when C4-AP was introduced near the H2A acidic patch. PPC formation by K MP is modulated by the NCP environment. Combined with the known intracellular formation of K MP, this study inspires investigating whether PPC formation by this NECM impacts cell function and viability.
DOI: 10.1021/jacs.6c03639
Authors: Zehui Zhou, Hanrui Yu, Marc M. Greenberg
Institution: Johns Hopkins University
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W7154711219