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Medientyp:
E-Artikel
Titel:
T Antigen Binds to Simian Virus 40 DNA at the Origin of Replication
Beteiligte:
Reed, Steven I.;
Ferguson, Jill;
Davis, Ronald W.;
Stark, George R.
Erschienen:
National Academy of Sciences of the United States of America, 1975
Erschienen in:Proceedings of the National Academy of Sciences of the United States of America
Sprache:
Englisch
ISSN:
0027-8424
Entstehung:
Anmerkungen:
Beschreibung:
<p>A technique employing ferritin-conjugated antibody has been developed to visualize specific protein-DNA complexes in the electron microscope and has been used to demonstrate the preferential binding of simian virus 40 (SV40) T antigen at or near the origin of replication of SV40 DNA, 0.67 fractional length clockwise from the EcoRI restriction endonuclease cleavage site. Purified covalently closed supercoiled circles of SV40 DNA were treated with partially purified T antigen and the complex was stabilized by crosslinking with glutaraldehyde. Hamster anti-T antigen γ -globulin, ferritin-labeled goat anti-hamster γ -globulin, and glutaraldehyde were then added sequentially. The location of the bound ferritin cores was measured with respect to the EcoRI cleavage site and the orientation of the cores relative to the ends of the DNA was determined with respect to the locations of Escherichia coli DNA unwinding protein, which binds to covalently closed supercoiled SV40 DNA at either of two preferred sites, 0.46 or 0.90 fractional length clockwise from the EcoRI cleavage site.</p>