Principal Research Interests

One of the major unsolved mysteries in molecular biology is the protein folding problem, how proteins adopt their three-dimensional, biologically active conformations. In particular, we are interested in understanding the mechanisms responsible for the formation of membrane spanning channels that transport proteins important for disease pathogenesis. Our laboratory focuses on two biological systems– the anthrax protective antigen from Bacillus anthracis, and CooC, a protein required for transport of CS1 pili from enterotoxigenic E. coli to the bacterial cell surface. We utilize a variety of biochemical and biophysical techniques, including circular dichroism, fluorescence and NMR to aid in our investigations.

The anthrax protective antigen is the B component of the anthrax toxin, an AB toxin secreted by B. anthracis that includes the enzymatic A components edema factor and lethal factor. Edema and lethal factor enter the cell cytosol through a pore formed by the protective antigen. The exact mechanism of pore formation is not understood, but is known to require the pH-dependent release of a receptor (anthrax toxin receptors 1 or 2) from within an endocytic vesicle. Understanding what initiates the release of the receptor and subsequent membrane insertion is of particular interest to our laboratory. Our laboratory has developed methods for biosynthetically incorporating 2- and 4-fluorohistidine into proteins. 2-fluorohistidine has a side-chain pKa of 1, and uniform labeling of the protective antigen with this amino acid has been used to address the role of histidine protonation in governing the conformational change from a prepore to a membrane spanning pore, as well as receptor release.

CS1 pili from enterotoxigenic E. coli are long hair-like fibers that extend from the bacterial surface and are required for adherence to host cells. The CS1 pilus assembly system utilizes only four proteins for proper assembly of a pilus – CooA, the major pilus subunit that comprises the polymeric fiber; CooB, a chaperone required for folding of CooA; CooC, an outer membrane usher protein required for assembly at the outer membrane; and CooD, the tip of the pilus which contains a lectin domain for interacting with host cell surfaces. How pili, including CS1, are assembled is not well understood. Of central importance is CooC, the membrane spanning usher protein required for transport of CS1 pili to the cell surface. Through our studies on the related Pap system from pylenophritic E. coli, CooC initiates the transfer of a chaperone subunit complex (CooB-CooA), to a subunit-subunit complex (CooA-CooA). Understanding how CooC facilitates this transfer process is a major focus of our research. Understanding the mechanism of CS1 pilus assembly and transport at the outer membrane may aid in the development of therapeutics that block pilus assembly or transport.
 

1. Wimalasena, D.S., Cramer, J., Janowiak, B.E., Juris, S.J., Melnyk, S.J. Anderson, D.E., Kirk, K.L., Collier, R.J., and Bann, J.G. Effect of 2-fluorohistidine labeling of the anthrax protective antigen on stability, pore formation and translocation. Biochemistry, 2007 (November epub).

2. Bann, J.G., Cegelski, L., and Hultgren, S.J. LRP6 Holds the Key to the Entry of Anthrax Toxin. Cell, 2006, 124: 1119.

3. Eichler, J.F., Cramer, J.C., Kirk, K.L., and Bann, J.G. Biosynthetic Incorporation of fluoro-histidine into proteins in E. coli: a new probe of macromolecular structure, ChemBioChem, 2005, 6: 2170-2173.

4. Bann, J.G., Pinkner, J.S., Frieden, C., and Hultgren, S.J. Catalysis of protein folding by chaperones in pathogenic bacteria, Proc. Natl. Acad. Sci. USA. 2004, 101: 17389.

5. Bann, J.G., Frieden, C. “Folding and domain-domain interactions of the chaperone PapD measured by 19F-NMR,” Biochemistry, 2004, 43: 13775.

6. Frieden, C., Hoeltzli, S.D., Bann, J.G. The preparation of 19F-labeled proteins for NMR studies, Methods in Enzymology, 2004, 180: 400.

7. Bann, J.G., Dodson, K.W., Frieden, C., and Hultgren, S.J. Adhesive Pili of the Chaperone-Usher Family. In: Escherichia coli: Virulence Mechanisms of a Versatile Pathogen, 2002. Ed. Michael Donnenberg. Academic Press.

8. Bann, J.G., Pinkner, J., Hultgren, S.J., and Frieden, C. Real-time and equilibrium 19F-NMR studies reveal the role of domain-domain interactions in the folding of the chaperone PapD Proc. Natl. Acad. Sci. USA. 2002, 99: 709.