Structural and functional analysis of the anti-malarial drug target prolyl-tRNA synthetase

Abstract

Aminoacyl-tRNA synthetases (aaRSs) drive protein translation in cells and hence these are essential enzymes across life. Inhibition of these enzymes can halt growth of an organism by stalling protein translation. Therefore, small molecule targeting of aaRS active sites is an attractive avenue from the perspective of developing anti-infectives. Febrifugine and its derivatives like halofuginone (HF) are known to inhibit prolyl-tRNA synthetase of malaria parasite Plasmodium falciparum. Here, we present functional and crystallographic data on P. falciparum prolyl-tRNA synthetase (PfPRS). Using immunofluorescence data, we show that PfPRS is exclusively resident in the parasite cytoplasm within asexual blood stage parasites. The inhibitor HF interacts strongly with PfPRS in a non-competitive binding mode in presence or absence of ATP analog. Intriguingly, the two monomers that constitute dimeric PfPRS display significantly different conformations in their active site regions. The structural analyses presented here provide a framework for development of febrifugine derivatives that can seed development of new anti-malarials.

Structure of Prolyl-tRNA Synthetase-Halofuginone Complex Provides Basis for Development of Drugs against Malaria and Toxoplasmosis

“…The structure was solved at 2.3 Å resolution by molecular replacement method using PfPRS-apo (PDB code 4TWA) as a search model (Figure 2A). The organization and overall fold of PfPRS ternary complex is similar to that in our earlier report of PfPRS-apo structure (root-mean-square deviation [RMSD] is 0.6 Å for 475 Ca atoms) (Jain et al, 2014). PfPRS shares $54% identity with human PRS and yielded an RMSD of 1.3 Å for 470 Ca atoms, with limited deviations in AB binding and Z domains only ( Figure 2B).…”

“…The 36-member P. falciparum parasite tRNA synthetase family, which together governs transmission of genetic information into proteins, has been focus of intense research lately (Bhatt et al, 2009(Bhatt et al, , 2011Istvan et al, 2011;Jackson et al, 2011;Jain et al, 2014;Khan et al, 2011Khan et al, , 2013aKhan et al, , 2013bKhan et al, , 2014Koh et al, 2012;Pham et al, 2014). The recent discovery and validation of cladosporin utility against multiple malaria parasitic stages, and its specificity for PfKRS, has been a seminal contribution in validating aaRSs as new drug targets (Hoepfner et al, 2012;Khan et al, 2013aKhan et al, , 2014.…”

“…These findings generated intense excitement about the molecular target of HF in Plasmodium parasites. We then showed direct and strong HF binding to the Plasmodium falciparum PRS (PfPRS), thus establishing the cellular target of this drug in malaria parasites (Jain et al, 2014). In the past two decades, many synthetic derivatives of FF and HF have been reported with potent anti-malarial activity (Kikuchi et al, 2006, and it is very likely that these compounds work by targeting the malaria protein translation machinery by inhibition of the PRS.…”

“…However, lack of experimentally verified ''hit'' compounds has hampered the discovery of potent malaria parasite inhibitors against these protein targets (Chandra et al, 2005;Gill et al, 2009;Navadgi et al, 2006;Sharma and Sharma, 2011;Sharma et al, 2008). Encouragingly, several groups have been investigating the structure-function attributes of aaRSs within malaria parasites (Bhatt et al, 2009(Bhatt et al, , 2011Istvan et al, 2011;Jackson et al, 2011;Jain et al, 2014;Khan et al, 2011Khan et al, , 2013aKhan et al, , 2013bKhan et al, , 2014Koh et al, 2012;Pham et al, 2014). Targeting parasitic aaRSs can provide an additional drug component in current multidrug anti-malarial therapy (Bhatt et al, 2009(Bhatt et al, , 2011Hoepfner et al, 2012;Istvan et al, 2011;Jackson et al, 2011;Jain et al, 2014;Khan et al, 2011Khan et al, , 2013aKhan et al, , 2013bKhan et al, , 2014Koh et al, 2012;Pham et al, 2014).…”

Combining traditional medicine and modern chemistry to fight malaria

“…Modeling of the interaction of halofuginone with the prolyl-tRNA synthetases from S. cerevisiae and P. falciparum, structures of which have recently been solved (8,9), revealed a potential molecular mechanism of the binding of the drug to its target, and insight into why the drug only inhibits the P. falciparum enzyme but not the yeast enzyme.…”