Structural characterization of OM proteins from Gram-negative pathogens

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Our structures of Y. pestis FyuA and pesticin allowed us to engineer a novel phage therapy drug against a Gram-negative pathogen: We solved the structure of FyuA, a TonB-dependent iron transporter required for virulence in bubonic plague, with and without its cognate siderophore, ferric yersiniabactin. At the same time, we determined the structure of a bacteriocin called pesticin that uses FyuA to cross the outer membrane. Once inside the periplasm, pesticin kills the cell by degrading the peptidoglycan layer. From our structure we discovered that the killing domain of pesticin resembles phage T4 lysozyme, so we engineered a hybrid bacterial-phage toxin that contains a bacterial targeting domain (to FyuA) and a phage killing domain. We showed that the hybrid lysine evades the natural protection mechanism of toxin-producing strains and kills all Yersinia strains tested, both in vitro and in vivo (mouse model of bubonic plague). This is the first demonstration of phage therapy for Gram-negative pathogens because until now, no one knew how to transport the toxin across the outer membrane. This work was recently published at PNAS* and has received considerable attention, including a Nature Microbiology Reviews Highlight. We have recently begun a project to identify and characterize outer membrane proteins that are either essential or required for virulence in a variety of Gram-negative pathogens, since these proteins may make good vaccine components and drug targets. Structural characterization of several candidate proteins is currently underway. *Lukacik, P., Barnard, T.J., Keller, P.W., Chaturvedi, K., Seddiki, N., Fairman, J.W., Noinaj, N., Kirby, T.L., Henderson, J.P., Steven, A.C., Hinnebusch, B.J. & Buchanan, S.K. (2012). Structural engineering of a phage lysin that targets Gram-negative pathogens. Proc. Natl. Acad. Sci. USA, 109, 9857-9862. PMCID: PMC3382549 Research Highlighted: In this Issue Proc. Natl. Acad. Sci. USA, 109, 9667-9668 (2012). Research Highlighted: Nat. Rev. Microbiol. 10, 520-521 (2012). Research Highlighted: SciBX 5(27); doi:10.1038/scibx.2012.706 (2012) (Nature publishing group).