Structure of the origin-binding domain of simian virus 40 large T antigen bound to DNA.
The large T antigen (T-ag) protein binds to and activates DNA replication from the origin of DNA replication (ori) in simian virus 40 (SV40). Here, we determined the crystal structures of the T-ag origin-binding domain (OBD) in apo form, and bound to either a 17 bp palindrome (sites 1 and 3) or a 23 bp ori DNA palindrome comprising all four GAGGC binding sites for OBD. The T-ag OBDs were shown to interact with the DNA through a loop comprising Ser147-Thr155 (A1 loop), a combination of a DNA-binding helix and loop (His203-Asn210), and Asn227. The A1 loop traveled back-and-forth along the major groove and accounted for most of the sequence-determining contacts with the DNA. Unexpectedly, in both T-ag-DNA structures, the T-ag OBDs bound DNA independently and did not make direct protein-protein contacts. The T-ag OBD was also captured bound to a non-consensus site ATGGC even in the presence of its canonical site GAGGC. Our observations taken together with the known biochemical and structural features of the T-ag-origin interaction suggest a model for origin unwinding.
Bochkareva
Elena
E
Banting and Best Department of Medical Research & Department of Medical Genetics and Microbiology, University of Toronto, 100 College St., Toronto, Ontario, Canada.
Martynowski
Dariusz
D
Seitova
Almagoul
A
Bochkarev
Alexey
A
D000595:Amino Acid Sequence; D000957:Antigens, Viral, Tumor / Q000737:chemistry* / Q000378:metabolism*; D001483:Base Sequence; D018360:Crystallography, X-Ray; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018741:Replication Origin; D013539:Simian virus 40 / Q000737:chemistry*; D014779:Virus Replication / Q000502:physiology
D000957:Antigens, Viral, Tumor; D004279:DNA, Viral