Reconstitution of Mammalian Enzymatic Deacylation Reactions in Live Bacteria Using Native Acylated Substrates

Emanuel M. Avrahami, Shahar Levi, Eyal Zajfman, Clil Regev, Oshrit Ben-David, Eyal Arbely

Research output: Contribution to journalArticlepeer-review

7 Scopus citations


Lysine deacetylases (KDACs) are enzymes that catalyze the hydrolysis of acyl groups from acyl-lysine residues. The recent identification of thousands of putative acylation sites, including specific acetylation sites, created an urgent need for biochemical methodologies aimed at better characterizing KDAC-substrate specificity and evaluating KDACs activity. To address this need, we utilized genetic code expansion technology to coexpress site-specifically acylated substrates with mammalian KDACs, and study substrate recognition and deacylase activity in live Escherichia coli. In this system the bacterial cell serves as a "biological test tube" in which the incubation of a single mammalian KDAC and a potential peptide or full-length acylated substrate transpires. We report novel deacetylation activities of Zn2+-dependent deacetylases and sirtuins in bacteria. We also measure the deacylation of propionyl-, butyryl-, and crotonyl-lysine, as well as novel deacetylation of Lys310-acetylated RelA by SIRT3, SIRT5, SIRT6, and HDAC8. This study highlights the importance of native interactions to KDAC-substrate recognition and deacylase activity.

Original languageEnglish
Pages (from-to)2348-2354
Number of pages7
JournalACS Synthetic Biology
Issue number10
StatePublished - 19 Oct 2018


  • KDAC
  • genetic code expansion
  • histone deacetylase
  • lysine acetylation
  • sirtuin

ASJC Scopus subject areas

  • Biomedical Engineering
  • Biochemistry, Genetics and Molecular Biology (miscellaneous)


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