SV40 large T antigen hexamer structure: domain organization and DNA-induced conformational changes.
Simian Virus 40 replication requires only one viral protein, the Large T antigen (T-ag), which acts as both an initiator of replication and as a replicative helicase (reviewed in ). We used electron microscopy to generate a three-dimensional reconstruction of the T-ag hexameric ring in the presence and absence of a synthetic replication fork to locate the T-ag domains, to examine structural changes in the T-ag hexamer associated with DNA binding, and to analyze the formation of double hexamers on and off DNA. We found that binding DNA to the T-ag hexamer induces large conformational changes in the N- and C-terminal domains of T-ag. Additionally, we observed a significant increase in density throughout the central channel of the hexameric ring upon DNA binding. We conclude that conformational changes in the T-ag hexamer are required to accommodate DNA and that the mode of DNA binding may be similar to that suggested for some other ring helicases. We also identified two conformations of T-ag double hexamers formed in the presence of forked DNA: with N-terminal hexamer-hexamer contacts, similar to those formed on origin DNA, or with C-terminal contacts, which are unlike any T-ag double hexamers reported previously.
VanLoock
Margaret S
MS
Department of Biochemistry and Molecular Genetics, University of Virginia Health Sciences, Box 800733, Charlottesville, VA 22908-0733, USA.
Alexandrov
Alexander
A
Yu
Xiong
X
Cozzarelli
Nicholas R
NR
Egelman
Edward H
EH
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000952:Antigens, Polyomavirus Transforming / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004265:DNA Helicases / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D008854:Microscopy, Electron; D008958:Models, Molecular; D011487:Protein Conformation; D017434:Protein Structure, Tertiary
D000952:Antigens, Polyomavirus Transforming; D004247:DNA; D000251:Adenosine Triphosphatases; D004265:DNA Helicases