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A Scintillation Proximity Assay for Poly(ADP-ribose) Polymerase

✍ Scribed by Anissa Cheung; Jie Zhang

Publisher
Elsevier Science
Year
2000
Tongue
English
Weight
199 KB
Volume
282
Category
Article
ISSN
0003-2697

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✦ Synopsis

Poly(ADP-ribose) polymerase (PARP) is an abundant nuclear protein in most of the eukaryotic tissues. When activated by DNA damage, PARP synthesizes poly(ADP-ribose) from NAD. Conventional radioactive PARP enzyme assay requires the separation of the polymer product from the NAD substrate, a rate-limiting step that hampers large-scale chemical library screening to identify novel small-molecule PARP inhibitors. By using biotinylated NAD, we have developed a scintillation proximity assay (SPA) for PARP. We demonstrated that PARP can incorporate the biotinylated ADP-ribose units into the radioactive poly-(ADP-ribose) polymer, which can directly bind and excite the streptavidin-conjugated scintillation beads. PARP-SPA can be readily adapted to a 96-well format for automatic high-throughput screening for PARP inhibitors.

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