Dataset Information

Structural and evolutionary analyses of the Plasmodium falciparum chloroquine resistance transporter.

ABSTRACT: Mutations in the Plasmodium falciparum chloroquine resistance transporter (PfCRT) confer resistance to several antimalarial drugs such as chloroquine (CQ) or piperaquine (PPQ), a partner molecule in current artemisinin-based combination therapies. As a member of the Drug/Metabolite Transporter (DMT) superfamily, the vacuolar transporter PfCRT may translocate substrate molecule(s) across the membrane of the digestive vacuole (DV), a lysosome-like organelle. However, the physiological substrate(s), the transport mechanism and the functional regions of PfCRT remain to be fully characterized. Here, we hypothesized that identification of evolutionary conserved sites in a tertiary structural context could help locate putative functional regions of PfCRT. Hence, site-specific substitution rates were estimated over Plasmodium evolution at each amino acid sites, and the PfCRT tertiary structure was predicted in both inward-facing (open-to-vacuole) and occluded states through homology modeling using DMT template structures sharing <15% sequence identity with PfCRT. We found that the vacuolar-half and membrane-spanning domain (and especially the transmembrane helix 9) of PfCRT were more conserved, supporting that its physiological substrate is expelled out of the parasite DV. In the PfCRT occluded state, some evolutionary conserved sites, including positions related to drug resistance mutations, participate in a putative binding pocket located at the core of the PfCRT membrane-spanning domain. Through structural comparison with experimentally-characterized DMT transporters, we identified several conserved PfCRT amino acid sites located in this pocket as robust candidates for mediating substrate transport. Finally, in silico mutagenesis revealed that drug resistance mutations caused drastic changes in the electrostatic potential of the transporter vacuolar entry and pocket, facilitating the escape of protonated CQ and PPQ from the parasite DV.

SUBMITTER: Coppee R

PROVIDER: S-EPMC7076037 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structural and evolutionary analyses of the Plasmodium falciparum chloroquine resistance transporter.

Coppée Romain R Sabbagh Audrey A Clain Jérôme J

Scientific reports 20200316 1

Mutations in the Plasmodium falciparum chloroquine resistance transporter (PfCRT) confer resistance to several antimalarial drugs such as chloroquine (CQ) or piperaquine (PPQ), a partner molecule in current artemisinin-based combination therapies. As a member of the Drug/Metabolite Transporter (DMT) superfamily, the vacuolar transporter PfCRT may translocate substrate molecule(s) across the membrane of the digestive vacuole (DV), a lysosome-like organelle. However, the physiological substrate(s) ...[more]

PMID: 32179795

Similar Datasets

Project description:BACKGROUND: In Haiti where chloroquine (CQ) is widely used for malaria treatment, reports of resistance are scarce. However, recent identification of CQ resistance genotypes in one site is suggestive of an emerging problem. Additional studies are needed to evaluate genetic mutations associated with CQ resistance, especially in the Plasmodium falciparum multi-drug resistance-1 gene (pfmdr1) while expanding the already available information on P. falciparum CQ transporter gene (pfcrt) in Haiti. METHODS: Blood samples were collected on Whatman filter cards (FTA) from eight clinics spread across Haiti. Following the confirmation of P. falciparum in the samples, PCR protocols were used to amplify regions of pfmdr1and pfcrt codons of interest, (86, 184, 1034, 1042, and 1246) and (72-76), respectively. Sequencing and site-specific restriction enzyme digestions were used to analyse these DNA fragments for the presence of single nucleotide polymorphisms (SNPs) known to confer resistance to anti-malarial drugs. RESULTS: P. falciparum infection was confirmed in160 samples by amplifying a segment of the P. falciparum 18S small subunit ribosomal RNA gene (pfssurrna). The sequence of pfmdr1 in 54 of these samples was determined between codons 86,184 codons 1034, 1042 and 1246. No sequence differences from that of the NF54 clone 3D7 were found among the 54 samples except at codon 184, where a non-silent mutation was found in all samples predicted to alter the amino acid sequence replacing tyrosine with phenylalanine (Y184F). This altered sequence was also confirmed by restriction enzyme digestion. The sequence of pfmdr1 at codons 86, 184, 1034 and 1042 encoded the NFSN haplotype. The sequence of pfcrt codons 72-76 from 79 samples was determined and found to encode CVMNK, consistent with a CQ sensitive genotype. CONCLUSION: The presence of the Y184F mutation in pfmdr1 of P. falciparum parasites in Haiti may have implications for resistance to antimalarial drugs. The absence of mutation in pfcrt at codon 76 among 79 isolates tested suggests that sensitivity to CQ in Haiti remains common. Wide-spread screening of the pfmdr1 and pfcrt especially among patients experiencing treatment failure may be a useful tool in early detection of the emergence of antimalarial drug resistance in Haiti.

Dataset Information

Structural and evolutionary analyses of the Plasmodium falciparum chloroquine resistance transporter.

Publications

Structural and evolutionary analyses of the Plasmodium falciparum chloroquine resistance transporter.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets