Poly (ADP-ribose) polymerases (PARPs)

PARP Enzymatic Assays

We use an ELISA-based method to detect the enzymatic activity of PARP family members. Histones are coated on the plate, and PARP proteins, together with biotin-labeled NAD, are added to the reaction. PARP1, PARP2, and PARP3 require single-strand DNA breaks for activation. As the reaction proceeds, PARP1 forms biotin-labeled ADP-ribose chains on histones. Streptavidin-labeled HRP and an ultrasensitive detection reagent are then used to generate the assay signal.

PARP Trapping Assay

When PARP forms a complex with DNA and NAD, it normally dissociates from DNA as ADP-ribose chains are generated. In this assay, biotin labels are introduced at single-stranded DNA breaks and PARP proteins are tagged with His tags. Inhibitor-induced PARP–DNA retention can then be detected through a fluorescence resonance energy transfer signal between donor and acceptor tags.

PARP Binding Assay

PARP1 and PARP2 binding can be evaluated using Fluorescence Polarization (FP) and Surface Plasmon Resonance (SPR). FP supports rapid, high-throughput assessment of small-molecule binding affinity, while SPR provides real-time kinetic analysis of association and dissociation. Together, these methods support comprehensive profiling of PARP binding properties.