Structure of the Yersinia pestis type III export complex

Project: Research project

Project Details

Description

DESCRIPTION (provided by applicant): The use of aerosolized Yersinia pestis as
a biological weapon could potentially produce massive numbers of casualties.
Weaponized plague could be rendered resistant to most antibiotics and
delivered via an aerosol. Thus, in order to protect the military and civilian
populations from both endemic and recombinant Y. pestis, new biological
countermeasures must be developed that target essential pathogen
vulnerabilities. The Y. pestis type III secretion (TTS) system enables the
bacterium to subvert or destroy eukaryotic cells via the delivery of anti-host
effector proteins. Disruption of the TTS process renders Y. pestis avirulent,
indicating that the TTS process represents an attractive target for
therapeutic intervention. Assembly of the TTS apparatus requires the
participation of at least 21 Ysc (Yersinis secretion) proteins, Ysc proteins
are hypothesized to assemble into a supramolecular structure that functions as
a protein secretion and injection device. We propose to investigate the
assembly and structure of the TTS apparatus. In Specific Aim 1, experiments
are described to identify the protein interactions involved in the assembly of
the TTS complex. Immunoprecipitation methodologies and yeast two- and three-
hybrid studies will identify Ysc proteins that directly interact with one another.
The overall objective of Specific Aim 2 is to establish conditions for the solubilization,
purification and visualization of the assembled or partially assembled TTS
complex. Environmental scanning electron microscopy (ESEM) of fixed samples
and/or transmission electron microscopy (TEM) of negatively-stained samples
will be used to visualize the surface-exposed and/or isolated TTS complex
and/or partially assembled intermediates of this complex. In Specific Aim 3,
experiments designed to identify and characterize chromosomally-encoded proteins
that are required for the assembly and function of the TTS complex are described.
Completion of these Specific Aims will not only provide essential information about the
assembly and function of the Y. pestis TTS complex, but may also facilitate
the development and/or design of novel therapeutics that target this essential
virulence system.
StatusFinished
Effective start/end date9/1/015/31/06

Funding

  • National Institute of Allergy and Infectious Diseases: $227,250.00
  • National Institute of Allergy and Infectious Diseases: $193,163.00
  • National Institute of Allergy and Infectious Diseases: $227,250.00
  • National Institute of Allergy and Infectious Diseases: $255,136.00

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