Protein kinase and ATP-binding activity associated with the 72-kdalton single-stranded DNA-binding protein from early region 2A of human adenovirus type 5.
We have developed monoclonal antibodies which react specifically with the human adenovirus type 5 early region 2A 72-kdalton phosphoprotein (72 kDa) and its 48-kdalton proteolytic cleavage product (48 kDa) and have used these antibodies to study a number of properties of these viral polyPeptides. Fluorescent antibody staining indicated that the 72 kDa was found almost entirely in the nucleus, generally in discreet patches. Preparations of the 72 kDa, purified by immunoprecipitation or by single-stranded DNA-cellulose column chromatography and incubated with [gamma-32P]ATP, were found to contain protein kinase activity. Using photoaffinity labelling with 8-azido-[alpha-32P]ATP, the 72 kDa was shown to be an ATP-binding protein. The ATP-binding site was probably in the amino terminal region because the 48 kDa which lacks approximately 120 residues at the amino terminus failed to bind ATP. Experiments carried out with immunoprecipitates from wild-type and temperature-sensitive (ts 125) infected cells suggested that the ts mutant-induced kinase activity was not more thermolabile than the wild-type protein. Thus although the present results indicated that protein kinase activity is associated with the 72 kDa, it is still unclear whether such activity is intrinsic to the 72 kDa or present in an enzyme associated with it. While it is probably not intrinsic to the carboxy terminal region affected by the ts 125 mutation, it is still possible that it resides in an amino terminal domain.
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000258:Adenovirus Infections, Human / Q000378:metabolism; D000260:Adenoviruses, Human / Q000378:metabolism*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006358:Hot Temperature; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D011233:Precipitin Tests; D011494:Protein Kinases / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D000255:Adenosine Triphosphate; D011494:Protein Kinases