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Journal of Bacteriology, May 1999, p. 2765-2772, Vol. 181, No. 9
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Role of CIS in Replication of an IncB Plasmid

J. Praszkier and A. J. Pittard*

Department of Microbiology, University of Melbourne, Parkville, Victoria 3052, Australia

Received 29 September 1998/Accepted 16 February 1999

Replication of the IncB plasmid pMU720 requires the synthesis of the cis-acting RepA protein and the presence of two DNA elements, ori and CIS. CIS is the 166-bp sequence separating the RepA coding sequence from ori. To investigate how this organization of the pMU720 replicon contributes to the mechanism of initiation of replication, mutations in the sequence and/or the length of CIS were introduced into the CIS region and their effects on the efficiency of replication of the pMU720 replicon in vivo was determined. The CIS region was found to be composed of two domains. The repA-proximal domain, which showed strong transcription termination activity, could be replaced by equivalent sequences from I-complex and IncL/M plasmids, whose replicons are organized in the same fashion as pMU720. Replacement by a trpA transcription terminator afforded only partial replication activity. The repA-distal domain was shown to be a spacer whose role was to position sequence(s) within ori on the correct face of the DNA helix vis-à-vis the repA-proximal portion of CIS. A model for the loading of RepA protein onto ori is discussed.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, University of Melbourne, Royal Parade, Parkville, Victoria 3052, Australia. Phone: 61 3 9344 5679. Fax: 61 3 9347 1540. E-mail: aj.pittard{at}microbiology.unimelb.edu.au.

Journal of Bacteriology, May 1999, p. 2765-2772, Vol. 181, No. 9
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Betteridge, T., Yang, J., Pittard, A. J., Praszkier, J. (2004). Role of RepA and DnaA Proteins in the Opening of the Origin of DNA Replication of an IncB Plasmid. J. Bacteriol. 186: 3785-3793 [Abstract] [Full Text]  
  • Odegrip, R., Coomber, D., Eldridge, B., Hederer, R., Kuhlman, P. A., Ullman, C., FitzGerald, K., McGregor, D. (2004). CIS display: In vitro selection of peptides from libraries of protein-DNA complexes. Proc. Natl. Acad. Sci. USA 101: 2806-2810 [Abstract] [Full Text]  
  • Betteridge, T., Yang, J., Pittard, A. J., Praszkier, J. (2003). Interaction of the Initiator Protein of an IncB Plasmid with Its Origin of DNA Replication. J. Bacteriol. 185: 2210-2218 [Abstract] [Full Text]  
  • Praszkier, J., Murthy, S., Pittard, A. J. (2000). Effect of CIS on Activity in trans of the Replication Initiator Protein of an IncB Plasmid. J. Bacteriol. 182: 3972-3980 [Abstract] [Full Text]  


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