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Essentiality of Plasmodium falciparum plasmepsin V.

ABSTRACT: The malaria parasite replicates within erythrocytes. The pathogenesis of clinical malaria is in large part due to the capacity of the parasite to remodel its host cell. To do this, intraerythrocytic stages of Plasmodium falciparum export more than 300 proteins that dramatically alter the morphology of the infected erythrocyte as well as its mechanical and adhesive properties. P. falciparum plasmepsin V (PfPMV) is an aspartic protease that processes proteins for export into the host erythrocyte and is thought to play a key role in parasite virulence and survival. However, although standard techniques for gene disruption as well as conditional protein knockdown have been previously attempted with the pfpmv gene, complete gene removal or knockdown was not achieved so direct genetic proof that PMV is an essential protein has not been established. Here we have used a conditional gene excision approach combining CRISPR-Cas9 gene editing and DiCre-mediated recombination to functionally inactivate the pfpmv gene. The resulting mutant parasites displayed a severe growth defect. Detailed phenotypic analysis showed that development of the mutant parasites was arrested early in the ring-to-trophozoite transition in the erythrocytic cycle following gene excision. Our findings are the first to elucidate the effects of PMV gene disruption, showing that it is essential for parasite viability in asexual blood stages. The mutant parasites can now be used as a platform to further dissect the Plasmodium protein export pathway.

SUBMITTER: Boonyalai N

PROVIDER: S-EPMC6281190 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Essentiality of Plasmodium falciparum plasmepsin V.

Boonyalai Nonlawat N Collins Christine R CR Hackett Fiona F Withers-Martinez Chrislaine C Blackman Michael J MJ

PloS one 20181205 12

The malaria parasite replicates within erythrocytes. The pathogenesis of clinical malaria is in large part due to the capacity of the parasite to remodel its host cell. To do this, intraerythrocytic stages of Plasmodium falciparum export more than 300 proteins that dramatically alter the morphology of the infected erythrocyte as well as its mechanical and adhesive properties. P. falciparum plasmepsin V (PfPMV) is an aspartic protease that processes proteins for export into the host erythrocyte a ...[more]

PMID: 30517136

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Project description:Malaria is a devastating disease, and its management is only achieved through chemotherapy. However, resistance to available medication is still a challenge; therefore, there is an urgent need for the discovery and development of therapeutics with a novel mechanism of action to counter the resistance scourge consistent with the currently available antimalarials. Recently, plasmepsin V was validated as a therapeutic target for the treatment of malaria. The pepsin-like aspartic protease anchored in the endoplasmic reticulum is responsible for the trafficking of parasite-derived proteins to the erythrocytic surface of the host cells. In this study, a small library of compounds was preliminarily screened in vitro to identify novel modulators of Plasmodium falciparum plasmepsin V (PfPMV). The results obtained revealed kaempferol, quercetin, and shikonin as possible PfPMV inhibitors, and these compounds were subsequently probed for their inhibitory potentials using in vitro and in silico methods. Kaempferol and shikonin noncompetitively and competitively inhibited the specific activity of PfPMV in vitro with IC50 values of 22.4 and 43.34 μM, respectively, relative to 62.6 μM obtained for pepstatin, a known aspartic protease inhibitor. Further insight into the structure-activity relationship of the compounds through a 100 ns molecular dynamic (MD) simulation showed that all the test compounds had a significant affinity for PfPMV, with quercetin (-36.56 kcal/mol) being the most prominent metabolite displaying comparable activity to pepstatin (-35.72 kcal/mol). This observation was further supported by the compactness and flexibility of the resulting complexes where the compounds do not compromise the structural integrity of PfPMV but rather stabilized and interacted with the active site amino acid residues critical to PfPMV modulation. Considering the findings in this study, quercetin, kaempferol, and shikonin could be proposed as novel aspartic protease inhibitors worthy of further investigation in the treatment of malaria.

Dataset Information

Essentiality of Plasmodium falciparum plasmepsin V.

Publications

Essentiality of Plasmodium falciparum plasmepsin V.

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