SV40 T antigen binds directly to the large subunit of purified DNA polymerase alpha.
Purified SV40 large T antigen and purified DNA polymerase alpha-primase form a complex detectable by ELISA and by a modified immunoblotting technique. The interaction is specific for the large catalytic subunit of polymerase alpha. The amino terminal 83 amino acids of T antigen are both necessary and sufficient for binding to the polymerase. However, antibody epitopes located in the carboxy terminal ATPase domain of T antigen are masked in the polymerase-T antigen complex, and complex formation is inhibited by an antibody directed against the carboxy terminus of T antigen, suggesting that this region of T antigen, though not required for binding, is in close proximity to the bound polymerase. The affinity of human DNA polymerase alpha for T antigen is approximately 10-fold greater than that of polymerase alpha from calf thymus, consistent with the interpretation that polymerase alpha is at least in part responsible for the primate-specific replication of SV40 DNA in vivo and in vitro. The results suggest that specific protein-protein interaction between DNA polymerase alpha and T antigen plays an important role in viral DNA replication.
Dornreiter
I
I
Institute for Biochemistry, Munich, FRG.
Höss
A
A
Arthur
A K
AK
Fanning
E
E
D000818:Animals; D000952:Antigens, Polyomavirus Transforming; D001665:Binding Sites; D002417:Cattle; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D004257:DNA Polymerase II / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D019915:DNA Primase; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis; D015151:Immunoblotting; D007700:Kinetics; D046911:Macromolecular Substances; D012316:RNA Nucleotidyltransferases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013539:Simian virus 40 / Q000276:immunology* / Q000378:metabolism; D013950:Thymus Gland / Q000201:enzymology
D000952:Antigens, Polyomavirus Transforming; D000939:Epitopes; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D019915:DNA Primase; D012316:RNA Nucleotidyltransferases; D004257:DNA Polymerase II