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Journal of Bacteriology, April 1999, p. 2572-2583, Vol. 181, No. 8
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Genetic Analysis of the Mobilization and Leading Regions of the IncN plasmids pKM101 and pCU1

E. Suzanne Paterson,1 Margret I. Moré,2 Gansen Pillay,2 Christina Cellini,2 Roger Woodgate,3 Graham C. Walker,4 V. N. Iyer,1 and Stephen C. Winans2,*

Department of Biology, Carleton University, Ottawa, Ontario, Canada K1S 5B61; Section of Microbiology, Cornell University, Ithaca, New York 148532; Biology Department, Massachusetts Institute of Technology, Cambridge, Massachusetts 021394; and National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892-27253

Received 20 October 1998/Accepted 4 February 1999

The conjugative IncN plasmids pKM101 and pCU1 have previously been shown to contain identical oriT sequences as well as conserved restriction endonuclease cleavage patterns within their tra regions. Complementation analysis and sequence data presented here indicate that these two plasmids encode essentially identical conjugal DNA-processing proteins. This region contains three genes, traI, traJ, and traK, transcribed in the same orientation from a promoter that probably lies within or near the conjugal transfer origin (oriT). Three corresponding proteins were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and complementation analysis confirmed that this region contains three tra complementation groups. All three proteins resemble proteins of the IncW plasmid R388 and other plasmids thought to have roles in processing of plasmid DNA during conjugation. The hydropathy profile of TraJ suggests a transmembrane topology similar to that of several homologous proteins. Both traK and traI were required for efficient interplasmid site-specific recombination at oriT, while traJ was not required. The leading region of pKM101 contains three genes (stbA, stbB, and stbC), null mutations in which cause elevated levels of plasmid instability. Plasmid instability was observed only in hosts that are proficient in interplasmid recombination, suggesting that this recombination can potentially lead to plasmid loss and that Stb proteins somehow overcome this, possibly via site-specific multimer resolution.

* Corresponding author. Mailing address: Department of Microbiology, Cornell University, Ithaca, NY 14853. Phone: (607) 255-2413. Fax: (607) 255-3904. E-mail: scw2{at}cornell.edu.

Journal of Bacteriology, April 1999, p. 2572-2583, Vol. 181, No. 8
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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