Real-time investigation of SV40 large T-antigen helicase activity using surface plasmon resonance.
The simian virus 40 (SV40) genome is a model system frequently employed for investigating eukaryotic replication. Large T-antigen (T-ag) is a viral protein responsible for unwinding the SV40 genome and recruiting necessary host factors prior to replication. In addition to duplex unwinding T-ag possesses G-quadruplex DNA helicase activity, the physiological consequence of which is unclear. However, formation of G-quadruplex DNA structures may be involved in genome maintenance and function, and helicase activity to resolve these structures may be necessary for efficient replication. We report the first real-time investigation of SV40 T-ag helicase activity using surface plasmon resonance (SPR). In the presence of ATP, T-ag was observed to bind to immobilized single-stranded DNA, forked duplex DNA, and the human telomeric foldover quadruplex DNA sequence. Inhibition of T-ag duplex helicase activity was observable in real-time and the intramolecular quadruplex was unwound.
Plyler
Jason
J
Department of Chemistry and Biochemistry, Texas State University-San Marcos, San Marcos, TX 78666, USA.
Jasheway
Karl
K
Tuesuwan
Bodin
B
Karr
Jessica
J
Brennan
Jarryd S
JS
Kerwin
Sean M
SM
David
Wendi M
WM
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000378:metabolism*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication / Q000235:genetics / Q000276:immunology; D004279:DNA, Viral / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D054856:G-Quadruplexes; D006801:Humans; D013539:Simian virus 40 / Q000201:enzymology* / Q000235:genetics / Q000276:immunology*; D020349:Surface Plasmon Resonance / Q000379:methods*
D000957:Antigens, Viral, Tumor; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D000251:Adenosine Triphosphatases; D004265:DNA Helicases