Simian virus 40 tumor antigen: isolation of the origin-specific DNA-binding domain.
To localize the origin-specific DNA-binding domain on the simian virus 40 tumor (T) antigen molecule, we used limited proteolysis with trypsin to generate fractional peptides for analysis. A 17,000-Mr peptide was found to be capable of binding not only to calf thymus DNA, but also specifically to the simian virus 40 origin of DNA replication. This approximately 130-amino-acid peptide was derived from the extreme N-terminus of the T antigen and represented less than one-fifth of the entire molecule. The coding sequence for this tryptic peptide was located approximately between 0.51 and 0.67 map units (excluding the intron, which maps between 0.54 and 0.59). Since the first 82 amino acids are shared between large T and small t antigens, and since the latter does not bind DNA, it can be concluded that the sequence between isoleucine 83 and approximately arginine 130 is necessary for origin-specific binding by the T antigen. We also observed that in vivo phosphorylation of the T antigen within this region completely abolished the ability of the 17,000-Mr peptide to bind DNA. This observation is consistent with the idea that DNA binding by the T antigen is regulated by posttranslational modifications.
D000818:Animals; D000956:Antigens, Viral / Q000302:isolation & purification* / Q000378:metabolism; D000957:Antigens, Viral, Tumor; D001665:Binding Sites; D002460:Cell Line; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity; D004247:DNA / Q000378:metabolism*; D004261:DNA Replication; D004587:Electrophoresis, Agar Gel; D007668:Kidney; D010455:Peptides / Q000302:isolation & purification / Q000378:metabolism; D013539:Simian virus 40 / Q000276:immunology*; D014357:Trypsin
D000956:Antigens, Viral; D000957:Antigens, Viral, Tumor; D010455:Peptides; D004247:DNA; D014357:Trypsin