Poster abstracts

Poster number 6 submitted by Paul Kelly

Developing a Novel Screen for Inhibitors of Leishmania major Aminoacyl-tRNA Synthetases

Paul Kelly (Molecular, Cellular, and Developmental Biology Program, The Ohio State University), Tammy Bullwinkle (Department of Microbiology, The Ohio State University), David Ardell (Program in Quantitative Systems Biology, University of California, Merced), Roger Linington (Department of Chemistry, Simon Fraser University), Abhay Satoskar (Department of Microbiology and the Department of Pathology, The Ohio State University), Michael Ibba (Department of Microbiology, The Ohio State University)

Abstract:
Leishmania major is a eukaryotic parasite that infects upwards of a million individuals a year. Current therapeutic options remain limited and have adverse effects on humans. While antibiotics are able to target prokaryotic cellular components specifically, developing therapies against eukaryotic pathogens is more challenging due to conservation between potential pathogen drug targets and their host counterparts. Previous studies identified anti-fungal therapies that target pathogen aminoacyl-tRNA synthetases (aaRS) with some degree of specificity, indicating that these essential enzymes could be developed as drug targets for a variety of infections. AaRS transfer amino acids on to their corresponding tRNA in a two-step process: 1) binding to the free cognate amino acid in an ATP-dependent manner (activation) and 2) aminoacylation of the amino acid on to the cognate tRNA (transfer). Using bioinformatic approaches, we have identified several aaRSs in L. major that appear to be potential targets for anti-leishmanial therapies. Here we have implemented a medium-throughput drug screen and tested for aminoacylation inhibitors against the L. major alanyl-tRNA synthetase (AlaRS). From the 120 compound-mixes tested, 3 mixes prevented Ala-tRNAAla aminoacylation. We further examined which step of the AlaRS activity the compounds may be inhibiting, and using radiolabeled ATP-PPi assays, show that the drugs impair amino acid activation. Furthermore, when we performed similar assays against the human AlaRS enzyme, there was no effect on activation. These in vitro results suggest that we may have identified anti-leishmanial compounds that would justify future pharmacokinetic studies in vivo.

Keywords: aminoacyl-tRNA synthetases, drug screen, Leishmania major