Bovine papillomavirus type 1 E1 ATPase activity does not depend on binding to DNA nor to viral E2 protein.
Replication of bovine papillomavirus type 1 (BPV-1) DNA has been shown to require two viral proteins known to interact in a molecular complex: E2, a transcription activator, and E1, another nuclear phosphoprotein, which binds to the replication origin and for which helicase/ATPase activities have previously been reported. Here we characterize the BPV-1 E1 ATPase activity. In contrast to Seo et al. (Proceedings of the National Academy of Sciences, USA, 90, 702-706, 1993), we were able to detect this activity in the absence of nucleic acid in partially purified preparations of either E1 protein or of E1-E2 protein complex. Measurements of specific activity and kinetic parameters gave similar values for preparations of various kinds. ATPase activity was quantitatively retained by immunoprecipitates obtained by using anti-E1 or, in the case of E1-E2 complex, anti-E2 antibodies. Significantly, preparations of bacterially expressed glutathione S-transferase-E1 fusion protein exhibited levels of DNA-independent ATPase activity comparable to those of baculovirus-expressed E1. The presence of nucleic acids of various types, including stoichiometric amounts of a BPV-1 ori DNA fragment containing E1 and E2 binding sites, did not grossly affect E1 ATPase activity, the most notable effect being a 2-fold stimulation by unspecific ssDNA. Altogether, our results indicate that BPV-1 E1 possesses an intrinsic ATPase activity which does not depend on the presence of nucleic acid; moreover, they render unlikely any modulation of E1 ATPase activity due to binding either E2 protein or target DNA sequences, or as a result of protein phosphorylation.
Santucci
S
S
INSERM U273, Centre de Biochimie, Université de Nice, France.
Bonne-Andréa
C
C
Clertant
P
P
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D052688:Bovine papillomavirus 1 / Q000201:enzymology*; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*
D004268:DNA-Binding Proteins; C069814:E1 protein, Bovine papillomavirus; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014764:Viral Proteins; D004247:DNA; D000251:Adenosine Triphosphatases