Exploring the Histone Acylome through Incorporation of γ-Thialysine on Histone Tails

Abstract

Histone lysine acetyltransferases (KATs) catalyze the transfer of the acetyl group from acetyl Coenzyme A to lysine residues in histones and non-histone proteins. Here, we report biomolecular studies on epigenetic acetylation and related acylation reactions of lysine and γ-thialysine, a cysteine-derived lysine mimic, which can be site-specifically introduced to histone peptides and histone proteins. Enzyme assays demonstrate that human KATs catalyze an efficient acetylation and propionylation of histone peptides that possess lysine and γ-thialysine. Enzyme kinetics analyses reveal that lysine-and γ-thialysinecontaining histone peptides exhibit indistinguishable K m values, whereas small differences in k cat values were observed. This work highlights that γ-thialysine may act as a representative and easily accessible lysine mimic for chemical and biochemical examinations of posttranslationally modified histones.

Effect of lysine side chain length on histone lysine acetyltransferase catalysis

“…Enzyme kinetics showed p300 catalytic efficiency towards a collection of Lys analogs; a synthetic H4K8 peptide was found to be the preferred substrate, followed by D-Lys and γ-thiahomolysine that showed very poor catalytic efficiency 48 . More recently, γ-thialysine was demonstrated to be efficiently accepted by a panel of human KATs for the transfer of acetyl, propionyl and butyryl moieties from their respective acyl CoA cosubstrates 49 . To provide a better insight into the role of lysine side chain on histone acetylation, this study is aimed at understanding of the relevance of lysine side chain length on human KAT catalysis.…”

Probing lysine posttranslational modifications by unnatural amino acids

“…107 The synthetically easily accessible lysine mimic γ-thialysine (K c ) was also investigated as a substrate for human KATs. 108 Interestingly, γ-thialysine behaved as an excellent analogue for acetylation, also accepting the slightly larger propionyl group, further establishing γ-thialysine as a suitable mimic for studies of PTMs of lysine with a potential application in full-length proteins. 108 Amide-derived lysine analogues were not accepted by KATs, while KMTs had the capacity of accepting the smallest change introduced here.…”

“…108 Interestingly, γ-thialysine behaved as an excellent analogue for acetylation, also accepting the slightly larger propionyl group, further establishing γ-thialysine as a suitable mimic for studies of PTMs of lysine with a potential application in full-length proteins. 108 Amide-derived lysine analogues were not accepted by KATs, while KMTs had the capacity of accepting the smallest change introduced here. 74 Notably, γ-difluorolysine was verified as an excellent lysine mimic for the KAT-catalysed acetylation reaction, providing a tool for studying lysine acetylation by 19 F NMR spectroscopy (Fig.…”

“…In the studies of KATs, a variety of analogues have been used to assess their substrate scope. 71,74,106–108 Human KATs appeared to be relatively promiscuous when it comes to the chain length of the lysine substrate; whereas shorter lysine analogues were not acetylated, extension of the side chain by three carbons was still tolerated in some cases, while a two-carbon extension was universally accepted (Fig. 7C, Chain length).…”

Comparison of Molecular Recognition of Trimethyllysine and Trimethylthialysine by Epigenetic Reader Proteins

“…Cysteine can be selectively alkylated to obtain analogues that mimic naturally occurring amino acids. Among these are arginine [18], lysine [19,20] and different posttranslationally modified variants of lysine, including acetylated [21] and methylated analogues [22]. For the purpose of generating methylated lysine, simple bromides can be used for chemoselective reaction with the cysteine thiol to obtain intact histone proteins that possess simplest methylated lysine analogues (MLAs) [23].…”