Activation of the ATPase activity of adeno-associated virus Rep68 and Rep78.
Rep68 and Rep78 DNA helicases, encoded by adeno-associated virus 2 (AAV2), are required for replication of AAV viral DNA in infected cells. They bind to imperfect palindromic elements in the inverted terminal repeat structures at the 3'- and 5'-ends of virion DNA. The ATPase activity of Rep68 and Rep78 is stimulated up to 10-fold by DNA containing the target sequence derived from the inverted terminal repeat; nontarget DNA stimulates ATPase activity at 50-fold higher concentrations. Activation of ATPase activity of Rep68 by DNA is cooperative with a Hill coefficient of 1.8 +/- 0.2. When examined by gel filtration at 0.5 M NaCl in the absence of DNA, Rep68 self-associates in a concentration-dependent manner. In the presence of DNA containing the binding element, Rep68 (and Rep78) forms protein-DNA complexes that exhibit concentration-dependent self-association in gel filtration analysis. The ATPase activity of the isolated Rep68-DNA and Rep78-DNA complexes is not activated by additional target DNA. Results of sedimentation velocity experiments in the presence of saturating target DNA are consistent with Rep68 forming a hexamer of the protein with two copies of the DNA element. Activation of the ATPase activity of Rep68 is associated with the formation of a protein-DNA oligomer.
Dignam
Susan S
SS
Department of Biochemistry and Cancer Biology, University of Toledo College of Medicine, 3035 Arlington Avenue, Toledo, Ohio 43614-5804, USA.
Correia
John J
JJ
Nada
Shadia E
SE
Trempe
James P
JP
Dignam
John David
JD
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D000229:Dependovirus / Q000201:enzymology*; D019281:Dimerization; D004789:Enzyme Activation; D007700:Kinetics; D014764:Viral Proteins / Q000378:metabolism*
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D014764:Viral Proteins; C068734:rep proteins, Adeno-associated virus 2; D000251:Adenosine Triphosphatases