Rapid Identification of Bacterial Virulence Factors.

摘要

The main objective of this work is to demonstrate that Rapid Virulence Factor Discovery System (RVFDS) can be used to identify virulence factors of MRSA and Brucella abortus. This will be accomplished by constructing an MRSA and B. abortus random expression library (REL) in E. coll. The MRSA REL contains hundreds of potential virulence factors. Putative virulence factors that interacted with the ECM components fibronectin, laminin, and plasminogen, collagen, and anti-MRSA polyclonal antibodies will be eluted from pull-down columns that contain these bait proteins. Putative MRSA virulence factors will be identified by LC-MS/MS. Additional virulence factors will be identified using a wider range of ECM components as bait proteins. In the Phase I investigation the average size of the REL insert was in the 1.5 to 2.0kb range. In Phase II the average size of the insert will be increased to 3.0kb in order to decrease redundancy. Larger inserts and REL protein sizes will also facilitate LC-MS/MS identification and shorten the time needed for data analysis. Potential proteins that are discovered in Phase II work may serve as targets for future vaccines, antimicrobials, and therapeutic measures.