Structure of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase–dihydropteroate synthase from Plasmodium vivax sheds light on drug resistance

Abstract

The genomes of the malaria-causing parasites encode a protein fused of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) and dihydropteroate synthase (DHPS) domains that catalyze sequential reactions in the folate biosynthetic pathway. Whereas higher organisms derive folate from their diet and lack the enzymes for its synthesis, most eubacteria and a number of lower eukaryotes including malaria parasites synthesize tetrahydrofolate via DHPS. () and () HPPK-DHPSs are currently targets of drugs like sulfadoxine (SDX). The SDX effectiveness as an antimalarial drug is increasingly diminished by the rise and spread of drug-resistant mutations. Here, we present the crystal structure of HPPK-DHPS in complex with four substrates/analogs, revealing the bifunctionalHPPK-DHPS architecture in an unprecedented state of enzymatic activation. SDX's effect on HPPK-DHPS is due to 4-amino benzoic acid (ABA) mimicry, and the HPPK-DHPS structure sheds light on the SDX-binding cavity, as well as on mutations that effect SDX potency. We mapped five dominant drug resistance mutations inHPPK-DHPS: S382A, A383G, K512E/D, A553G, and V585A, most of which occur individually or in clusters proximal to the ABA-binding site. We found that these resistance mutations subtly alter the intricate enzyme/ABA/SDX interactions such that DHPS affinity for ABA is diminished only moderately, but its affinity for SDX is changed substantially. In conclusion, theHPPK-DHPS structure rationalizes and unravels the structural bases for SDX resistance mutations and highlights architectural features in HPPK-DHPSs from malaria parasites that can form the basis for developing next-generation anti-folate agents to combat malaria parasites.

Plasmodium falciparum DHFR and DHPS Mutations Are Associated With HIV-1 Co-Infection and a Novel DHPS Mutation I504T Is Identified in Western Kenya

“…The 3D structure of P. falciparum DHPS was not available at the time this study was initiated but has recently become available ( Chitnumsub et al, 2019 ). Accordingly, we used a crystal structure of P. vivax DHPS in complex with substrates 6-hydroxymethyl-7,8-dihydropterin pyrophosphate (DHPPP) and 4-aminobenzoic acid ( p ABA) ( Yogavel et al, 2018 ) to build a homology model of P. falciparum DHPS, which we subsequently compared to the published structure ( Chitnumsub et al, 2019 ). The experimental structure of P. vivax DHPS in complex with DHPPP and p ABA substrates was downloaded from the Protein Data Bank (PDB) server (PDB ID: 5Z79) ( Manickam et al, 2018 ) and the structure prediction wizard from the PRIME module of Schrodinger suite was used for homology modeling ( Jacobson et al, 2002 ; Jacobson et al, 2004 ).…”

Profiles of Kelch mutations in Plasmodium falciparum across South Asia and their implications for tracking drug resistance

“…Aside from propelling basic research directed at identifying the molecular markers for ACT failure, the international community is also racing towards the identification of new drug targets and scaffolds to cover for the eventual emergency of complete ACT failure (Chhibber-Goel and Sharma, 2019; Dogovski et al, 2015; Jain et al, 2017; Jain, 2017; Khan et al, 2014; Yogavel et al, 2018a, Yogavel et al, 2018b). Over the past decade, numerous new drug targets have been identified, probed and then validated.…”

“…Over the past decade, numerous new drug targets have been identified, probed and then validated. Drug repurposing methodologies and structure-based conservation techniques that target invariant parasite house-keeping proteins may lead to novel foci for drug development (Dogovski et al, 2015; Jain et al, 2017; Jain V, 2017; Khan et al, 2014; Pazhayam et al, 2019; Yogavel et al, 2018a, Yogavel et al, 2018b). However, till then, surveillance and containment of mutant P. falciparum parasites that show decreased susceptibility to ART/ACT are of primary importance.…”

Genomic analysis of single nucleotide polymorphisms in malaria parasite drug targets

“…Structures of P . falciparum and P. vivax HPPK-DHPS are available in complex with substrates/analogues (PDB ID: 6JWW and 5Z79) [ 27 , 28 ]. Pf HPPK-DHPS is a dimer having HPPK and DHPS domains connected by a linker (Fig.…”