RESUMO
Plasmodium falciparum (Pf) prolyl-tRNA synthetase (ProRS) is one of the few chemical-genetically validated drug targets for malaria, yet highly selective inhibitors have not been described. In this paper, approximately 40,000 compounds were screened to identify compounds that selectively inhibit PfProRS enzyme activity versus Homo sapiens (Hs) ProRS. X-ray crystallography structures were solved for apo, as well as substrate- and inhibitor-bound forms of PfProRS. We identified two new inhibitors of PfProRS that bind outside the active site. These two allosteric inhibitors showed >100 times specificity for PfProRS compared to HsProRS, demonstrating this class of compounds could overcome the toxicity related to HsProRS inhibition by halofuginone and its analogues. Initial medicinal chemistry was performed on one of the two compounds, guided by the cocrystallography of the compound with PfProRS, and the results can instruct future medicinal chemistry work to optimize these promising new leads for drug development against malaria.
Assuntos
Aminoacil-tRNA Sintetases/antagonistas & inibidores , Antimaláricos/farmacologia , Inibidores Enzimáticos/farmacologia , Plasmodium falciparum/enzimologia , Sítios de Ligação , Clonagem Molecular , Descoberta de Drogas , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Modelos Moleculares , Plasmodium falciparum/efeitos dos fármacos , Conformação Proteica , Bibliotecas de Moléculas PequenasRESUMO
Burkholderia pseudomallei is a soil-dwelling bacterium endemic to Southeast Asia and Northern Australia. Burkholderia is responsible for melioidosis, a serious infection of the skin. The enzyme 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase (PGAM) catalyzes the interconversion of 3-phosphoglycerate and 2-phosphoglycerate, a key step in the glycolytic pathway. As such it is an extensively studied enzyme and X-ray crystal structures of PGAM enzymes from multiple species have been elucidated. Vanadate is a phosphate mimic that is a powerful tool for studying enzymatic mechanisms in phosphoryl-transfer enzymes such as phosphoglycerate mutase. However, to date no X-ray crystal structures of phosphoglycerate mutase have been solved with vanadate acting as a substrate mimic. Here, two vanadate complexes together with an ensemble of substrate and fragment-bound structures that provide a comprehensive picture of the function of the Burkholderia enzyme are reported.