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Journal of Bacteriology, June 2004, p. 3785-3793, Vol. 186, No. 12
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.12.3785-3793.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Role of RepA and DnaA Proteins in the Opening of the Origin of DNA Replication of an IncB Plasmid

T. Betteridge, J. Yang, A. J. Pittard, and J. Praszkier*

Department of Microbiology and Immunology, The University of Melbourne, Victoria 3010, Australia

Received 8 December 2003/ Accepted 1 March 2004

The replication initiator protein RepA of the IncB plasmid pMU720 was shown to induce localized unwinding of its cognate origin of replication in vitro. DnaA, the initiator protein of Escherichia coli, was unable to induce localized unwinding of this origin of replication on its own but enhanced the opening generated by RepA. The opened region lies immediately downstream of the last of the three binding sites for RepA (RepA boxes) and covers one turn of DNA helix. A 6-mer sequence, 5'-TCTTAA-3', which lies within the opened region, was essential for the localized unwinding of the origin in vitro and origin activity in vivo. In addition, efficient unwinding of the origin of replication of pMU720 in vitro required the native positioning of the binding sites for the initiator proteins. Interestingly, binding of RepA to RepA box 1, which is essential for origin activity, was not required for the localized opening of the origin in vitro.


* Corresponding author. Mailing address: Department of Microbiology and Immunology, The University of Melbourne, Victoria 3010, Australia. Phone: 61 3 8344 7751. Fax: 61 3 9347 1540. E-mail: judy{at}ariel.its.unimelb.edu.au.


Journal of Bacteriology, June 2004, p. 3785-3793, Vol. 186, No. 12
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.12.3785-3793.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.




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