The semisynthesis of site-specifically modified histones and histone-based probes of chromatin-modifying enzymes

Abstract

Histone post-translational modifications (PTMs) on lysine residues, including methylation, ubiquitylation, and sumoylation, have been studied using semisynthetic histones reconstituted into nucleosomes. These studies have revealed the in vitro effects of histone PTMs on chromatin structure, gene transcription, and biochemical crosstalk. However, the dynamic and transient nature of most enzyme-chromatin interactions poses a challenge toward identifying specific enzyme-substrate interactions. To address this, we report methodology for the synthesis of two ubiquitylated activity-based probe histones, H2BK120ub(G76C) and H2BK120ub(G76Dha), that may be used to trap enzyme active-site cysteines as disulfides or in the form of thioether linkages, respectively. The general synthetic method we report for converting ubiquitylated nucleosomes into activity-based probes may also be applied to other histone sites of ubiquitylation in order to facilitate the identification of enzyme-chromatin interactions.

Keywords: Activity-based probe; Histone; Nucleosome; Post-translational modifications; Semisynthesis; Ubiquitylation.