Abstract 1132 High-throughput Virtual Screening of Potential Novel Inhibitors for Enoyl Acyl Protein Carrier Reductase (FabV) in Yersinia pestis

Plague outbreaks have persisted throughout the 21st century, specifically in medically underserved regions. New strains of Yersinia pestis, the causative pathogen, have been identified as resistant to traditional antibacterial treatments such as triclosan and isoniazid. Novel compounds which inhibit the same pathway that these drugs target may serve as alternative treatments for this disease. FabV, an enoyl-acyl protein carrier reductase, is an essential enzyme in the fatty acid biosynthesis pathway (FAS-II) of many pathogenic bacteria, including Y. pestis. Since this metabolic pathway is essential for the organism's viability, and the enzyme does not have a human homologue, it is an attractive candidate for small molecule inhibitors as therapeutic agents. High scoring ligands that may serve as novel inhibitors were identified in a high-throughput virtual screening approach using GOLD, a molecular docking software. Previously studied and crystallized ligands were used as positive controls, alongside randomly chosen, but properties-matched negative controls, in order to validate the approach for this structure. Libraries containing thousands of ligands were then docked against the YpFabV crystal structure (PDB: 4BKR) directed to the NADH pocket. The native ligand found naturally in the crystal structure of YpFabV, NADH, yielded a GOLD score of 85.5. This score was used as a threshold to find top scoring compounds from ZINC, NIH, and ChemBridge Diversity Libraries. Top scoring ligands included SAM__37 (GOLD Score: 103.55) and SAM__79 (GOLD Score: 102.48) from the NIH Clinical Collection 3D Library. In order to validate these predictions, YpFabV was cloned into a pET expression plasmid and expressed in SOLUBL21 E. coli cells in order to maximize protein yield via lactose-inducible autoinduction. The protein was purified using nickel affinity chromatography. The product was characterized through SDS-PAGE and a pure protein of 44 kDa was verified. The melting temperature of the apoenzyme was determined through differential scanning fluorimetry (DSF). Top-scoring ligands' effects on the thermostability of the protein were then assayed using this method. Given these preliminary results, these compounds show promise as potential inhibitors of FabV, and may provide a line of novel therapeutics against the plague.