Model for T-antigen-dependent melting of the simian virus 40 core origin based on studies of the interaction of the beta-hairpin with DNA.
The interaction of simian virus 40 (SV40) T antigen (T-ag) with the viral origin has served as a model for studies of site-specific recognition of a eukaryotic replication origin and the mechanism of DNA unwinding. These studies have revealed that a motif termed the "beta-hairpin" is necessary for assembly of T-ag on the SV40 origin. Herein it is demonstrated that residues at the tip of the "beta-hairpin" are needed to melt the origin-flanking regions and that the T-ag helicase domain selectively assembles around one of the newly generated single strands in a manner that accounts for its 3'-to-5' helicase activity. Furthermore, T-ags mutated at the tip of the "beta-hairpin" are defective for oligomerization on duplex DNA; however, they can assemble on hybrid duplex DNA or single-stranded DNA (ssDNA) substrates provided the strand containing the 3' extension is present. Collectively, these experiments indicate that residues at the tip of the beta-hairpin generate ssDNA in the core origin and that the ssDNA is essential for subsequent oligomerization events.
Kumar
Anuradha
A
Department of Biochemistry A703, Tufts University School of Medicine, 136 Harrison Avenue, Boston, MA 02111, USA.
Meinke
Gretchen
G
Reese
Danielle K
DK
Moine
Stephanie
S
Phelan
Paul J
PJ
Fradet-Turcotte
Amélie
A
Archambault
Jacques
J
Bohm
Andrew
A
Bullock
Peter A
PA
D020816:Amino Acid Motifs / Q000235:genetics; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000378:metabolism*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication / Q000235:genetics / Q000502:physiology*; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D009841:Oligonucleotides; D018741:Replication Origin / Q000235:genetics*; D013539:Simian virus 40 / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
D000957:Antigens, Viral, Tumor; D004277:DNA, Single-Stranded; D009841:Oligonucleotides; D004265:DNA Helicases