The use of biotinylated poly(ADP-ribose) for studies on poly(ADP-ribose)-protein interaction.
Poly(ADP-ribose) is routinely detected by the use of radioactive polymers formed from labeled substrates. In this report a simple and time-saving method for the biotinylation and the detection of poly(ADP-ribose) on blots is described. The polymer modified by light-induced reaction with photobiotin was colorimetrically detected and quantified, using streptavidine-alkaline phosphatase conjugates. The separation of poly(ADP-ribose) chains on polyacrylamide gels was not affected by the biotinylation of the polymers. When biotinylated poly(ADP-ribose) was used to detect the poly(ADP-ribose) binding capability of proteins in ligand blots, the results were comparable to those obtained with poly([32P]ADP-ribose). Experiments with histones and rat liver nuclear proteins demonstrate that in studies on poly(ADP-ribose)-protein interaction, this method is applicable to the detection of poly(ADP-ribose) binding proteins.
Institute of Tumorbiology--Cancer Research, University of Vienna, Austria.
F M
FM
Narendja
G
G
Sauermann
D000345:Affinity Labels; D000469:Alkaline Phosphatase; D000818:Animals; D001345:Autoradiography / Q000379:methods; D001386:Azides; D001426:Bacterial Proteins; D001710:Biotin / Q000031:analogs & derivatives; D002467:Cell Nucleus / Q000378:metabolism; D003124:Colorimetry / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D007202:Indicators and Reagents; D007700:Kinetics; D008024:Ligands; D008027:Light; D009243:NAD / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D011064:Poly Adenosine Diphosphate Ribose / Q000302:isolation & purification / Q000378:metabolism*; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D019809:Streptavidin
D000345:Affinity Labels; D001386:Azides; D001426:Bacterial Proteins; D007202:Indicators and Reagents; D008024:Ligands; D010761:Phosphorus Radioisotopes; D011506:Proteins; C090641:biotinyl-poly(adenosine 5'-diphospho-5'-ribose); D009243:NAD; D011064:Poly Adenosine Diphosphate Ribose; D001710:Biotin; D019809:Streptavidin; C045549:photobiotin; D000469:Alkaline Phosphatase