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Journal of Bacteriology, March 2004, p. 1606-1613, Vol. 186, No. 6
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.6.1606-1613.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

A Subassembly of R27-Encoded Transfer Proteins Is Dependent on TrhC Nucleoside Triphosphate-Binding Motifs for Function but Not Formation

Matthew W. Gilmour and Diane E. Taylor*

Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta T6G 2R3, Canada

Received 1 December 2003/ Accepted 3 December 2003

The transfer of plasmid DNA molecules between bacterial cells is achieved by a large array of conjugative transfer proteins which assemble into both cytoplasmic and membrane-associated complexes. TrhC is a membrane-associated protein that is required for the transfer of the IncHI1 resistance plasmid R27. Homologous proteins are encoded in all known conjugative systems, and each contains characteristic nucleoside triphosphate (NTP)-binding domains. An assembly of R27-encoded proteins was previously visualized by use of a TrhC-green fluorescent protein fusion, which appeared as discrete membrane-associated fluorescent foci. We have utilized this experimental system to determine the requirements for assembly of this TrhC-associated protein complex, and we found that 12 of the other 18 R27 transfer proteins are required for focus formation. An individual focus possibly represents a subassembly comprised of some or all of these transfer proteins. These data support the notion that the transfer apparatus is a multicomponent structure. In contrast, substitutions and deletions within TrhC NTP-binding motifs had minor effects on focus formation, but these mutations did affect plasmid transfer and bacteriophage susceptibility. These results indicate that TrhC requires intact NTP-binding motifs to function during conjugative transfer but that these motifs are not essential for the assembly of TrhC into a complex with other transfer proteins.

* Corresponding author. Mailing address: Department of Medical Microbiology and Immunology, 1-28 Medical Sciences Building, University of Alberta, Edmonton, Alberta T6G 2R3 Canada. Phone: (780) 492-4777. Fax: (780) 492-7521. E-mail: diane.taylor{at}ualberta.ca.

Journal of Bacteriology, March 2004, p. 1606-1613, Vol. 186, No. 6
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.6.1606-1613.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Chen, I., Christie, P. J., Dubnau, D. (2005). The Ins and Outs of DNA Transfer in Bacteria. Science 310: 1456-1460 [Abstract] [Full Text]  
  • Gunton, J. E., Gilmour, M. W., Alonso, G., Taylor, D. E. (2005). Subcellular localization and functional domains of the coupling protein, TraG, from IncHI1 plasmid R27. Microbiology 151: 3549-3561 [Abstract] [Full Text]  


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