Expanded Genetic Code Technologies for Incorporating Modified Lysine at Multiple Sites

Post-translational lysine N-modifications (such as acetylation, methylation, crotonylation, butyrylation, propionylation, ubiquitylation and sumoylation) of specific lysine residues in proteins regulate a variety of biological processes, including transcription, protein degradation, apoptosis, nuclear transport and the cell cycle (Figure 1A). Lysine acetylation and methylation of histones, for example, are important for defining the epigenetic status and controlling transcriptional activation/repression, DNA repair, X-chromosome inactivation and genome imprinting, through alterations of chromatin structures (Figure 1B and C). Acetylation and methylation of lysine residues have also been increasingly discovered in non-histone proteins, including cytoskeletal proteins, molecular chaperones, nuclear import factors, ribosomal proteins, transcription factors and translation factors. The acetylation and deacetylation of lysine residues are catalyzed by acetyltransferases and deacetylases, respectively. Acetylated lysine residues are specifically recognised by bromodomain-containing proteins, which contribute to the maintenance of the transcriptional activity. On the other hand, the amounts of the mono-, diand trimethylated forms of lysine residues are balanced by the methyltransferase and demethylase activities. However, little is known about whether the three possible methylation statuses of the lysine residues cause differences in protein functions, and if so, how they exert their effects. Histones containing trimethylated lysine residues are enriched in promoters, transcription start sites, exons and the coding regions of active and inactive genes and are involved in transcriptional activation/repression. In contrast, histones containing monomethylated lysine residues are often enriched in enhancer regions, Figure 1. Post-translational modifications of histone proteins. A) Examples of naturally occurring N-modified lysine residues (Me3Lys, Ac-Lys, Prop-Lys, Crot-Lys and Buty-Lys) are shown. B) Representation of histone tails and their post-translational modifications. This figure was adapted from refs. [41e] and [46a,b] . C) Structure of a mononucleosome core particle (PDB ID: 1AOI). H2A (magenta and blue), H2B (orange and yellow), H3 (pink and green), H4 (grey and turquoise) and a-satellite DNA (orange, double-strand) are shown as ribbon models.