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Journal of Virology, September 2003, p. 9809-9816, Vol. 77, No. 18
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.18.9809-9816.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

The Cap Region of Topoisomerase I Binds to Sites near Both Ends of Simian Virus 40 T Antigen

Rupa Roy,1 Pamela Trowbridge,1 Zheng Yang,2 James J. Champoux,2 and Daniel T. Simmons1*

Department of Biological Sciences, University of Delaware, Newark, Delaware 19716-2590,1 Department of Microbiology, School of Medicine, University of Washington, Seattle, Washington 98195-72422

Received 1 May 2003/ Accepted 24 June 2003

Two independent binding sites on simian virus 40 (SV40) T antigen for topoisomerase I (topo I) were identified. One was mapped to the N-terminal domain (residues 83 to 160) by a combination of enzyme-linked immunosorbent assays (ELISAs) and glutathione S-transferase (GST) pull-down assays performed with various T antigen deletion mutants. The second was mapped to the C-terminal domain (residues 602 to 708). The region in human topo I that binds to both sites in T antigen was identified by ELISAs, GST pull-down assays, and double-hexamer binding assays with topo I deletion mutants. This region corresponds to a distinct domain on topo I known as the cap region that maps from residues 175 to 433. By combining these data with information about the structure of T-antigen double hexamers associated with origin DNA, we propose that the cap region of topo I associates specifically with both ends of the double hexamer bound to the SV40 origin to initiate DNA replication.

* Corresponding author. Mailing address: Department of Biological Sciences, University of Delaware, Newark, DE 19716-2590. Phone: (302) 831-8547. Fax: (302) 831-2281. E-mail: dsimmons{at}udel.edu.

Journal of Virology, September 2003, p. 9809-9816, Vol. 77, No. 18
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.18.9809-9816.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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