Installation of Trimethyllysine Analogs on Intact Histones via Cysteine Alkylation

Site-specific incorporation of post-translationally modified amino acids into proteins, including histones, has been a subject of great interest for chemical and biochemical communities. Here, we describe a site-specific incorporation of structurally simplest trimethyllysine analogs into position 4 of the intact histone H3 protein. An efficient alkylation of cysteine 4 of the recombinantly expressed histone H3 provides a panel of trimethyllysine analogs that differ in charge, charge density, sterics, and chain length. We demonstrate that H3 histone that bears trimethyllysine analogs can be further assembled into the octameric histone complex that constitutes the nucleosome. Binding studies showed that H3 histone that possesses trimethyllysine analogs is well recognized by a PHD3 reader domain of human JARID1A. This work provides important (bio)chemical tools for fundamental biomolecular studies aimed at unravelling the molecular basis of the higher order nucleosome and chromatin assemblies.