PARP Substrates and Biomarkers

摘要

PARylated proteins are enriched by treating cell lysates comprising PARylated proteins and DNA/RNA with an endonuclease that cleaves the DNA/RNA but not the PAR; and separating the PARylated proteins from the cleaved DNA/RNA. PARylation sites are labeled by eluting PARylated proteins from a PAR-affinity substrate with a nucleophilic amine exchange reactant, wherein the reactant labels PARylation sites of the proteins. Specific binding agents are identified by screening compounds for specific binding to a PARylated protein disclosed herein; and identifying one of the compounds as a specific binder of the protein. Antibodies which specifically bind PARylation sites are also disclosed.

主权项

1. A method for detecting sites of ADP-ribosylation comprising steps:
(a) enriching for PARylated proteins comprising treating cell lysates comprising PARylated proteins and DNA/RNA with an endonuclease that cleaves the DNA/RNA but not the PAR; and separating the PARylated proteins from the cleaved DNA/RNA; or (b) labeling PARylation sites comprising eluting PARylated proteins from a PAR-affinity substrate with a nucleophilic amine exchange reactant, wherein the reactant labels PARylation sites of the proteins.