Heterologous expression of the Treponema pallidum laminin-binding adhesin Tp0751 in the culturable spirochete Treponema phagedenis

Department of Biochemistry and Microbiology, University of Victoria, Victoria, BC, Canada V8W 3P6; Department of Medicine, University of Washington, Seattle, WA, USA 98195; Department of Oral Biology, SUNY at Buffalo, Buffalo, NY, USA 14214

^* To whom correspondence should be addressed. Email: caroc{at}uvic.ca.

	Abstract

Treponema pallidum subsp. pallidum, the causative agent of syphilis, is an unculturable, genetically intractable bacterium. Here we report the use of the shuttle vector pKMR4PEMCS for expression of a previously identified T. pallidum laminin-binding adhesin, Tp0751, in the non-adherent, culturable spirochete Treponema phagedenis. Heterologous expression of Tp0751 in T. phagedenis was confirmed via reverse transcriptase polymerase chain reaction analysis using tp0751-specific primers and immunofluorescence analysis using Tp0751-specific antibodies; the latter assay verified expression of the laminin-binding adhesin on the treponemal surface. Expression of Tp0751 within T. phagedenis was functionally confirmed via laminin-attachment assays, in which heterologous Tp0751 expression conferred upon T. phagedenis the capacity to attach to laminin. Further, specific inhibition of attachment of T. phagedenis heterologously expressing Tp0751 to laminin was achieved using purified antibodies raised against recombinant T. pallidum Tp0751. This is the first report of heterologous expression of a gene from an unculturable treponeme in T. phagedenis. This novel methodology will significantly advance the field of syphilis research by allowing targeted investigations of T. pallidum proteins purported to play a role in pathogenesis, and specifically host cell attachment, in the non-adherent spirochete T. phagedenis.