ELTA: Enzymatic Labeling of Terminal ADP-Ribose

Yoshinari Ando, Elad Elkayam, Robert Lyle McPherson, Morgan Dasovich, Shang Jung Cheng, Jim Voorneveld, Dmitri V. Filippov, Shao En Ong, Leemor Joshua-Tor, Anthony K.L. Leung

Research output: Contribution to journalArticlepeer-review

Abstract

ADP-ribosylation refers to the addition of one or more ADP-ribose groups onto proteins. The attached ADP-ribose monomers or polymers, commonly known as poly(ADP-ribose) (PAR), modulate the activities of the modified substrates or their binding affinities to other proteins. However, progress in this area is hindered by a lack of tools to investigate this protein modification. Here, we describe a new method named ELTA (enzymatic labeling of terminal ADP-ribose) for labeling free or protein-conjugated ADP-ribose monomers and polymers at their 2′-OH termini using the enzyme OAS1 and dATP. When coupled with various dATP analogs (e.g., radioactive, fluorescent, affinity tags), ELTA can be used to explore PAR biology with techniques routinely used to investigate DNA or RNA function. We demonstrate that ELTA enables the biophysical measurements of protein binding to PAR of a defined length, detection of PAR length from proteins and cells, and enrichment of sub-femtomole amounts of ADP-ribosylated peptides from cell lysates. Ando et al. describe a simple, efficient, and versatile platform technology called ELTA to label free or protein-conjugated ADP-ribose monomers and polymers with dATP analogs (radioactive, fluorescent, biotin, clickable tags, etc.). With these functionalized tags, ELTA simplifies the measurement, detection, and enrichment of various forms of ADP-ribose.

Original languageEnglish (US)
Pages (from-to)845-856.e5
JournalMolecular cell
Volume73
Issue number4
DOIs
StatePublished - Feb 21 2019

Keywords

  • ADP-ribose
  • ADP-ribosylated protein
  • ADP-ribosylation
  • ADP-ribosyltransferase
  • enzymatic labeling
  • mono(ADP-ribosyl)ated protein
  • oligoadenylate synthetase
  • poly(ADP-ribose)
  • poly(ADP-ribose) polymerase
  • poly(ADP-ribosyl)ated protein

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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