Purification of polyoma virus medium-size tumor antigen by immunoaffinity chromatography.
We have used antibodies against the synthetic peptide Lys-Arg-Ser-Arg-His-Phe, corresponding to the six COOH-terminal amino acids of the polyoma virus medium tumor (T) antigen, to purify the medium T antigen by affinity chromatography. Release of the medium T antigen from the anti-peptide antibody was achieved under mild conditions by using a large excess of the peptide in an isotonic buffer at neutral pH containing mixed detergents. This procedure yielded a 2,500-fold purification of the medium T antigen in a single step. The protein kinase activity associated with the medium T antigen was also released and was studied in this active state in solution. Sedimentation analysis showed that the bulk of the purified medium T antigen was in a monomeric form (Mr about 42,000) not associated with protein kinase activity. A small fraction of the medium T antigen was found in a rapidly sedimenting form (Mr about 200,000) that possessed protein kinase activity.
D000818:Animals; D000906:Antibodies; D000936:Antigen-Antibody Complex; D000951:Antigens, Neoplasm / Q000302:isolation & purification*; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D051379:Mice; D008970:Molecular Weight; D011120:Polyomavirus / Q000201:enzymology / Q000276:immunology*; D011494:Protein Kinases / Q000302:isolation & purification
D000906:Antibodies; D000936:Antigen-Antibody Complex; D000951:Antigens, Neoplasm; D011494:Protein Kinases