Azalysine analogues as probes for protein lysine deacetylation and demethylation

J Am Chem Soc. 2012 Mar 21;134(11):5138-48. doi: 10.1021/ja209574z. Epub 2012 Mar 12.


Reversible lysine acetylation and methylation regulate the function of a wide variety of proteins, including histones. Here, we have synthesized azalysine-containing peptides in acetylated and unacetylated forms as chemical probes of the histone deacetylases (HDAC8, Sir2Tm, and SIRT1) and the histone demethylase, LSD1. We have shown that the acetyl-azalysine modification is a fairly efficient substrate for the sirtuins, but a weaker substrate for HDAC8, a classical HDAC. In addition to deacetylation by sirtuins, the acetyl-azalysine analogue generates a novel ADP-ribose adduct that was characterized by mass spectrometry, Western blot analysis, and nuclear magnetic resonance spectroscopy. This peptide-ADP-ribose adduct is proposed to correspond to a derailed reaction intermediate, providing unique evidence for the direct 2'-hydroxyl attack on the O-alkylimidate intermediate that is formed in the course of sirtuin catalyzed deacetylation. An unacetylated azalysine-containing H3 peptide proved to be a potent inhibitor of the LSD1 demethylase, forming an FAD adduct characteristic of previously reported related structures, providing a new chemical probe for mechanistic analysis.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Acetylation
  • Aza Compounds / chemistry
  • Aza Compounds / metabolism*
  • Fluorescent Dyes / chemical synthesis
  • Fluorescent Dyes / chemistry
  • Fluorescent Dyes / metabolism*
  • Histone Deacetylases / chemistry
  • Histone Deacetylases / metabolism*
  • Histone Demethylases / chemistry
  • Histone Demethylases / metabolism*
  • Lysine / analogs & derivatives
  • Lysine / chemistry
  • Lysine / metabolism*
  • Methylation
  • Molecular Structure
  • Peptides / chemical synthesis
  • Peptides / chemistry
  • Peptides / metabolism*


  • Aza Compounds
  • Fluorescent Dyes
  • Peptides
  • Histone Demethylases
  • Histone Deacetylases
  • Lysine