Article Metrics


Online attention

Synthesis of Dimeric ADP-Ribose and Its Structure with Human Poly(ADP-ribose) Glycohydrolase

DOI: 10.1021/ja512528p DOI Help
PMID: 25706250 PMID Help

Authors: Michael J. Lambrecht (University of Illinois) , Matthew Brichacek (University of Illinois) , Eva Barkauskaite (University of Oxford) , Antonio Ariza (University of Oxford) , Ivan Ahel (University of Oxford) , Paul J. Hergenrother (University of Illinois)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of The American Chemical Society , VOL 137 (10) , PAGES 3558 - 3564

State: Published (Approved)
Published: March 2015
Diamond Proposal Number(s): 9306

Open Access Open Access

Abstract: Poly(ADP-ribosyl)ation is a common post-translational modification that mediates a wide variety of cellular processes including DNA damage repair, chromatin regulation, transcription, and apoptosis. The difficulty associated with accessing poly(ADP-ribose) (PAR) in a homogeneous form has been an impediment to understanding the interactions of PAR with poly(ADP-ribose) glycohydrolase (PARG) and other binding proteins. Here we describe the chemical synthesis of the ADP-ribose dimer, and we use this compound to obtain the first human PARG substrate-enzyme cocrystal structure. Chemical synthesis of PAR is an attractive alternative to traditional enzymatic synthesis and fractionation, allowing access to products such as dimeric ADP-ribose, which has been detected but never isolated from natural sources. Additionally, we describe the synthesis of an alkynylated dimer and demonstrate that this compound can be used to synthesize PAR probes including biotin and fluorophore-labeled compounds. The fluorescently labeled ADP-ribose dimer was then utilized in a general fluorescence polarization-based PAR-protein binding assay. Finally, we use intermediates of our synthesis to access various PAR fragments, and evaluation of these compounds as substrates for PARG reveals the minimal features for substrate recognition and enzymatic cleavage. Homogeneous PAR oligomers and unnatural variants produced from chemical synthesis will allow for further detailed structural and biochemical studies on the interaction of PAR with its many protein binding partners.

Subject Areas: Biology and Bio-materials

Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)

Discipline Tags:

Technical Tags: