Development of Antibacterials Targeting the MEP Pathway of Select Agents
Abstract
Over the duration of this award, five specific aims were pursued: 1) Express, purify, and characterize recombinant Y. pestis IspC and IspD, 2) Optimize HTS assay conditions for IspC and IspD, 3) Provide purified recombinant F. tularensis IspC and IspD for crystallization and structure determination, 4) Provide purified recombinant Y. pestis IspC and IspD protein for crystallization and structure determination, and 5) Evaluate structure-activity relationships of rationally designed inhibitor molecules in enzyme-based assays. Accordingly, we successfully cloned, expressed, purified, and enzymatically characterized the Y. pestis IspC. Expression difficulties with recombinant Y. pestis IspD prompted us to work instead with cloned F. tularensis IspD. We successfully established HTS conditions for both IspC and IspD assays. Pilot scale screening of molecular libraries has identified hit compounds for IspC and IspD, including a novel inhibitor of IspC that binds to an allosteric site on the enzyme. We have provided purified protein on-demand for protein crystallography, and we have thoroughly evaluated structure-activity relationships of several rationally designed inhibitors of IspC and IspD via enzyme-based assays.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jun 01, 2018
- Accession Number
- AD1063865
Entities
People
- Robin Couch
Organizations
- George Mason University