Discovering New Classes of Brugia malayi Asparaginyl-tRNA Synthetase Inhibitors and Relating Specificity to Conformational Change

Abstract

SLIDE software, which models the flexibility of protein and ligand side chains while docking, was used to screen several large databases to identify inhibitors of Brugia malayi asparaginyl-tRNA synthetase (AsnRS), a target for anti-parasitic drug design. Seven classes of compounds identified by SLIDE were confirmed as micromolar inhibitors of the enzyme. Analogs of one of these classes of inhibitors, the long side-chain variolins, cannot bind to the adenosyl pocket of the closed conformation of AsnRS due to steric clashes, though the short side-chain variolins identified by SLIDE apparently bind isosterically with adenosine. We hypothesized that an open conformation of the motif 2 loop also permits the long side-chain variolins to bind in the adenosine pocket and that their selectivity for Brugia relative to human AsnRS can be explained by differences in the sequence and conformation of this loop. Loop flexibility sampling using Rigidity Optimized Conformational Kinetics (ROCK) confirms this possibility, while scoring of the relative affinities of the different ligands by SLIDE correlates well with the compounds' ranks in inhibition assays. Combining ROCK and SLIDE provides a promising approach for exploiting conformational flexibility in structure-based screening and design of species selective inhibitors.

Scoring ligand similarity in structure‐based virtual screening

“…SLIDE is a computational tool which can efficiently screen hundreds of thousands of molecules to identify feasible ligand candidates with good steric and chemical complementarity to a chosen site on a protein target Kuhn, 1999, 2000). The realistic modeling of protein side-chain and ligand flexibility and the improved representation of the binding site by knowledge-based template design (Zavodszky et al, 2002;Tonero et al, in preparation) allow discrimination between true ligands and non-specific compounds and facilitate the discovery of new inhibitors (Sukuru et al, 2006). Presented here are novel thrombin inhibitors discovered by screening the Plated Compounds Database of the National Cancer Institute (NCI).…”

Structural basis for the enantiospecificities of R‐ and S‐specific phenoxypropionate/α‐ketoglutarate dioxygenases

“…Previous homology models have led to successes in elucidating or designing specificity‐conferring interactions in ligands. For instance, homology modeling of a cercarial (human parasite) elastase led to the development of an effective elastase inhibitor (Cohen et al 1991) and to understanding the specificity determinants for ligands binding to a parasite tRNA synthetase versus its human homolog (Sukuru et al 2006). Here, we test by site‐directed mutagenesis and kinetic analysis the residues predicted to be involved in substrate binding, enantiospecificity, or catalysis.…”

Aminoacyl-tRNA synthetases as drug targets in eukaryotic parasites

“…From this docking, 45 compounds were tested for their inhibition of the Bm AsnRS, as assayed by a modified malachite green assay, which provides a readout for the first step in aminoacylation reaction. Of the compounds screened, a handful inhibited AsnRS aminoacylation at mid-micromolar IC 50 s (Table 1) (Sukuru et al, 2006). Subsequent publication of a more detailed structure for the Brugia AsnRS in complex with a substrate analogue that acts as a competitive inhibitor (Crepin et al, 2011) may provide additional information to refine future docking experiments or to rationally improve existing inhibitors.…”