Escherichia coli and Salmonella spp. are a significant cause of human disease. An important virulence factor for these bacteria is curli. Curli are stable proteinacious fibers that contribute to biofilm formation, host colonization, immune activation and cell invasion. Curli also represent a compelling example of a naturally occurring amyloid fiber. Several important human diseases result from proteins that misfold into amyloid fibers. Curli assembly does not result from protein misfolding, but from a dedicated biogenesis pathway, providing a new paradigm for examining amyloidogenesis. Our long-term goal is to elucidate the molecular mechanism of curli biogenesis so that therapeutic protocols can be designed to attenuate their formation. We developed a testable model of curli assembly that suggests the major curli subunit protein CsgA is secreted from the cell as an unstructured protein that subsequently folds into a ?-sheet rich fiber on the cell surface. We hypothesize that the CsgB nucleator protein catalyzes CsgA's initial folding, but then fiber formation proceeds as a self-perpetuating process with the growing fiber tip able to serve as a template for CsgA polymerization. Secretion of CsgA and CsgB to the cell surface is dependent on the outer membrane localized CsgG protein. E. coli provides us a sophisticated genetic and biochemical system to explore the biogenesis of these unique fibers.
In Aim 1 the hypothesis that CsgB presents an amyloid-like template to CsgA during nucleation will be tested.
In Aim 2 the sequences in CsgA that facilitate its interaction with CsgB and drive its polymerization into an amyloid fiber will be determined. Finally, in Aim 3 we will test the hypothesis that CsgG forms a curli-specific secretion pore in the outer membrane. Collectively, the experiments described within this proposal will reveal the molecular and structural basis for CsgA secretion, nucleation and polymerization into a fiber.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI073847-05
Application #
8038270
Study Section
Prokaryotic Cell and Molecular Biology Study Section (PCMB)
Program Officer
Baqar, Shahida
Project Start
2007-04-01
Project End
2013-09-04
Budget Start
2011-04-01
Budget End
2013-09-04
Support Year
5
Fiscal Year
2011
Total Cost
$348,232
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
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