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Journal of Bacteriology, March 2004, p. 1415-1422, Vol. 186, No. 5
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.5.1415-1422.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

VirB1 Orthologs from Brucella suis and pKM101 Complement Defects of the Lytic Transglycosylase Required for Efficient Type IV Secretion from Agrobacterium tumefaciens

Christoph Höppner,1 Zhenying Liu,2 Natalie Domke,1 Andrew N. Binns,2 and Christian Baron1,3*

Bereich Mikrobiologie, Department Biologie I, Ludwig-Maximilians-Universität München, D-80638 Munich, Germany,1 Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6018,2 Department of Biology, McMaster University, Hamilton, Ontario L8S 4K1, Canada3

Received 24 July 2003/ Accepted 13 November 2003

Type IV secretion systems mediate conjugative plasmid transfer as well as the translocation of virulence factors from various gram-negative pathogens to eukaryotic host cells. The translocation apparatus consists of 9 to 12 components, and the components from different organisms are believed to have similar functions. However, orthologs to proteins of the prototypical type IV system, VirB of Agrobacterium tumefaciens, typically share only 15 to 30% identical amino acids, and functional complementation between components of different type IV secretion systems has not been achieved. We here report a heterologous complementation in the case of A. tumefaciens virB1 defects with its orthologs from Brucella suis (VirB1s) and the IncN plasmid pKM101 (TraL). In contrast, expression of the genes encoding the VirB1 orthologs from the IncF plasmid (open reading frame 169) and from the Helicobacter pylori cag pathogenicity island (HP0523) did not complement VirB1 functions. The complementation of VirB1 activity was assessed by T-pilus formation, by tumor formation on wounded plants, by IncQ plasmid transfer, and by IncQ plasmid recipient assay. Replacement of the key active-site Glu residue by Ala abolished the complementation by VirB1 from B. suis and by TraL, demonstrating that heterologous complementation requires an intact lytic transglycosylase active site. In contrast, the VirB1 active-site mutant from A. tumefaciens retained considerable residual activity in various activity assays, implying that this protein exerts additional effects during the type IV secretion process.

* Corresponding author. Mailing address: Department of Biology, McMaster University, Life Sciences Bldg., 1280 Main St. West, Hamilton, Ontario L8S 4K1, Canada. Phone: (905) 525-9140, ext. 26692. Fax: (905) 522-6066. E-mail: baronc{at}mcmaster.ca.

Journal of Bacteriology, March 2004, p. 1415-1422, Vol. 186, No. 5
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.5.1415-1422.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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