This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Smelkova, N. V.
Right arrow Articles by Borowiec, J. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Smelkova, N. V.
Right arrow Articles by Borowiec, J. A.

 Previous Article  |  Next Article 

Journal of Virology, November 1998, p. 8676-8681, Vol. 72, No. 11
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Synthetic DNA Replication Bubbles Bound and Unwound with Twofold Symmetry by a Simian Virus 40 T-Antigen Double Hexamer

Natalia V. Smelkovadagger and James A. Borowiec*

Department of Biochemistry and Kaplan Comprehensive Cancer Center, New York University Medical Center, New York, New York 10016

Received 28 May 1998/Accepted 27 July 1998

Dimerization of simian virus 40 T-antigen hexamers (TAgH) into double hexamers (TAgDH) on model DNA replication forks has been found to greatly stimulate T-antigen DNA helicase activity. To explore the interaction of TAgDH with DNA during unwinding, we examined the binding of TAgDH to synthetic DNA replication bubbles. Tests of replication bubble substrates containing different single-stranded DNA (ssDNA) lengths indicated that efficient formation of a TAgDH requires >= 40 nucleotides (nt) of ssDNA. DNase I probing of a substrate containing a 60-nt ssDNA bubble complexed with a TAgDH revealed that T antigen bound the substrate with twofold symmetry. The strongest protection was observed over the 5' junction on each strand, with 5 bp of duplex DNA and ~17 nt of adjacent ssDNA protected from nuclease cleavage. Stimulation of the T-antigen DNA helicase activity by an increase in ATP concentration caused the protection to extend in the 5' direction into the duplex region, while resulting in no significant changes to the 3' edge of strongest protection. Our data indicate that each TAgH encircles one ssDNA strand, with a different strand bound at each junction. The process of DNA unwinding results in each TAgH interacting with a greater length of DNA than was initially bound, suggesting the generation of a more highly processive helicase complex.

* Corresponding author. Mailing address: Department of Biochemistry and Kaplan Comprehensive Cancer Center, New York University Medical Center, 550 First Ave., New York, NY 10016. Phone: (212) 263-8453. Fax: (212) 263-8166. E-mail: borowj01{at}mcrcr.med.nyu.edu.

dagger Present address: Dept. of Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021.

Journal of Virology, November 1998, p. 8676-8681, Vol. 72, No. 11
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Castella, S., Burgin, D., Sanders, C. M. (2006). Role of ATP hydrolysis in the DNA translocase activity of the bovine papillomavirus (BPV-1) E1 helicase. Nucleic Acids Res 34: 3731-3741 [Abstract] [Full Text]  
  • Fletcher, R. J., Shen, J., Gomez-Llorente, Y., Martin, C. S., Carazo, J. M., Chen, X. S. (2005). Double Hexamer Disruption and Biochemical Activities of Methanobacterium thermoautotrophicum MCM. J. Biol. Chem. 280: 42405-42410 [Abstract] [Full Text]  
  • You, Z., Masai, H. (2005). DNA binding and helicase actions of mouse MCM4/6/7 helicase. Nucleic Acids Res 33: 3033-3047 [Abstract] [Full Text]  
  • Sclafani, Robert. A., Fletcher, R. J., Chen, X. S. (2004). Two heads are better than one: regulation of DNA replication by hexameric helicases. Genes Dev. 18: 2039-2045 [Full Text]  
  • Jiao, J., Simmons, D. T. (2003). Nonspecific Double-Stranded DNA Binding Activity of Simian Virus 40 Large T Antigen Is Involved in Melting and Unwinding of the Origin. J. Virol. 77: 12720-12728 [Abstract] [Full Text]  
  • Grainge, I., Scaife, S., Wigley, D. B. (2003). Biochemical analysis of components of the pre-replication complex of Archaeoglobus fulgidus. Nucleic Acids Res 31: 4888-4898 [Abstract] [Full Text]  
  • Alexandrov, A. I., Botchan, M. R., Cozzarelli, N. R. (2002). Characterization of Simian Virus 40 T-antigen Double Hexamers Bound to a Replication Fork. THE ACTIVE FORM OF THE HELICASE. J. Biol. Chem. 277: 44886-44897 [Abstract] [Full Text]  
  • Uhlmann-Schiffler, H., Seinsoth, S., Stahl, H. (2002). Preformed hexamers of SV40 T antigen are active in RNA and origin-DNA unwinding. Nucleic Acids Res 30: 3192-3201 [Abstract] [Full Text]  
  • Ott, R. D., Wang, Y., Fanning, E. (2002). Mutational Analysis of Simian Virus 40 T-Antigen Primosome Activities in Viral DNA Replication. J. Virol. 76: 5121-5130 [Abstract] [Full Text]  
  • Wu, C., Roy, R., Simmons, D. T. (2001). Role of Single-Stranded DNA Binding Activity of T Antigen in Simian Virus 40 DNA Replication. J. Virol. 75: 2839-2847 [Abstract] [Full Text]  
  • Chen, G., Stenlund, A. (2001). The E1 Initiator Recognizes Multiple Overlapping Sites in the Papillomavirus Origin of DNA Replication. J. Virol. 75: 292-302 [Abstract] [Full Text]  
  • Gai, D., Roy, R., Wu, C., Simmons, D. T. (2000). Topoisomerase I Associates Specifically with Simian Virus 40 Large-T-Antigen Double Hexamer-Origin Complexes. J. Virol. 74: 5224-5232 [Abstract] [Full Text]  
  • Weisshart, K., Taneja, P., Jenne, A., Herbig, U., Simmons, D. T., Fanning, E. (1999). Two Regions of Simian Virus 40 T Antigen Determine Cooperativity of Double-Hexamer Assembly on the Viral Origin of DNA Replication and Promote Hexamer Interactions during Bidirectional Origin DNA Unwinding. J. Virol. 73: 2201-2211 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»