<i>Plasmodium falciparum</i>
            cysteine protease falcipain-1 is not essential in erythrocytic stage malaria parasites

Sijwali, Puran Singh; Kato, Koichiro; Seydel, Karl B.; Gut, Jiří; Lehman, Julie; Klemba, Michael; Goldberg, Daniel E.; Miller, Louis H.; Rosenthal, Philip J.

doi:10.1073/pnas.0402738101

Cited by 121 publications

(77 citation statements)

References 38 publications

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“…Importantly, the FP2A knockout parasites were 50-fold more sensitive to pepstatin, an aspartic protease inhibitor [20]. The development of FP1 knockout parasite lines was independently reported by two groups demonstrating that this cysteine protease is not required for parasite invasion and growth in erythrocytes [21,22]. These results were in contrast to an earlier report showing that the FP1 enzyme plays an essential role in the merozoite invasion of erythrocytes [23].…”

Section: Introductioncontrasting

confidence: 56%

BDA-410: A novel synthetic calpain inhibitor active against blood stage malaria

Chen

Jeong

et al. 2007

Molecular and Biochemical Parasitology

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Falcipains, the papain-family cysteine proteases of the Plasmodium falciparum, are potential drug targets for malaria parasite. Pharmacological inhibition of falcipains can block the hydrolysis of hemoglobin, parasite development, and egress, suggesting that falcipains play a key role at the blood stage of parasite life cycle. In the present study, we evaluated the anti-malarial effects of BDA-410, a novel cysteine protease inhibitor as a potential antimalarial drug. Recombinant falcipain (MBP-FP-2B) and Plasmodium falciparum trophozoite extract containing native falcipains were used for enzyme inhibition studies in vitro. The effect of BDA-410 on the malaria parasite development in vitro as well as its anti-malarial activity in vivo was evaluated using the Plasmodium chabaudi infection rodent model. The 50% inhibitory concentrations of BDA-410 were determined to be 628 nM and 534 nM for recombinant falcipain-2B and parasite extract, respectively. BDA-410 inhibited the malaria parasite growth in vitro with an IC50 value of 173 nM causing irreversible damage to the intracellular parasite. In vivo, the BDA-410 delayed the progression of malaria infection significantly using a mouse model of malaria pathogenesis. The characterization of BDA-410 as a potent inhibitor of Plasmodium falciparum cysteine proteases, and the demonstration of its efficacy in blocking parasite growth both in vitro and in vivo assays identifies BDA-410 is an important lead compound for the development of novel anti-malarial drugs.

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Section: Introductioncontrasting

confidence: 56%

BDA-410: A novel synthetic calpain inhibitor active against blood stage malaria

Chen

Jeong

et al. 2007

Molecular and Biochemical Parasitology

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“…Gene disruption methods have been used to explore the function of these enzymes and their role in hemoglobin digestion. Disruption of the gene encoding falcipain 2 did not result in complete ablation of cysteine endopeptidase activity but did impair hemoglobin digestion and parasite growth (37), whereas disruption of falcipain 1 had no effect on either (38). Disruption of the genes encoding falcipain 3 (37) and dipeptidyl aminopeptidase (12) is lethal to malaria parasites, and hence these peptidases are considered critical to survival within the erythrocyte.…”

Section: Discussionmentioning

confidence: 99%

Overexpression of Leucyl Aminopeptidase in Plasmodium falciparum Parasites

Gardiner

Trenholme

Skinner‐Adams

et al. 2006

Journal of Biological Chemistry

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Malaria aminopeptidases are important in the generation and regulation of free amino acids that are used in protein anabolism and for maintaining osmotic stability within the infected erythrocyte. The intraerythrocytic development of malaria parasites is blocked when the activity of aminopeptidases is specifically inhibited by reagents such as bestatin. One of the major aminopeptidases of malaria parasites is a leucyl aminopeptidase of the M17 family. We reasoned that, when this enzyme was the target of bestatin inhibition, its overexpression in malaria cells would lead to a reduced sensitivity to the inhibitor. To address this supposition, transgenic Plasmodium falciparum parasites overexpressing the leucyl aminopeptidase were generated by transfection with a plasmid that housed the full-length gene. Transgenic parasites expressed a 65-kDa protein close to the predicted molecule size of 67.831 kDa for the introduced leucyl aminopeptidase, and immunofluorescence studies localized the protein to the cytosol, the location of the native enzyme. The product of the transgene was shown to be functionally active with cytosolic extracts of transgenic parasites exhibiting twice the leucyl aminopeptidase activity compared with wildtype parasites. In vitro inhibitor sensitivity assays demonstrated that the transgenic parasites were more resistant to bestatin (EC 50 64 M) compared with the parent parasites (EC 50 25 M). Overexpression of genes in malaria parasites would have general application in the identification and validation of targets for antimalarial drugs.

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“…Disruption of falcipain-3 could not be achieved, but the gene was readily replaced with a tagged functional copy, indicating that falcipain-3 is essential for erythrocytic parasites (14). The roles of falcipain-2Ј (a nearidentical copy of falcipain-2) and falcipain-1 are unknown; knock out of these two proteases led to no apparent alterations in erythrocytic parasites (14,15).…”

Section: Malaria Causes Hundreds Of Millions Of Illnesses Annually Andmentioning

confidence: 99%

Falcipain Cysteine Proteases Require Bipartite Motifs for Trafficking to the Plasmodium falciparum Food Vacuole

Subramanian

Sijwali

Rosenthal

2007

Journal of Biological Chemistry

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The Plasmodium falciparum cysteine proteases falcipain-2 and falcipain-3 hydrolyze hemoglobin in an acidic food vacuole to provide amino acids for erythrocytic malaria parasites. Trafficking to the food vacuole has not been well characterized. To study trafficking of falcipains, which include large membranespanning prodomains, we utilized chimeras with portions of the proteases fused to green fluorescent protein. The prodomains of falcipain-2 and falcipain-3 were sufficient to target green fluorescent protein to the food vacuole. Using serial truncations, deletions, and point mutations, we showed that both a 20-amino acid stretch of the lumenal portion and a 10-amino acid stretch of the cytoplasmic portion of the falcipain-2 prodomain were required for efficient food vacuolar trafficking. Mutants with altered trafficking were arrested at the plasma membrane, implicating trafficking via this structure. Our results indicate that falcipains utilize a previously undescribed bipartite motifdependent mechanism for targeting to a hydrolytic organelle, suggesting inhibition of this unique mechanism as a new means of antimalarial chemotherapy.

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Plasmodium falciparum cysteine protease falcipain-1 is not essential in erythrocytic stage malaria parasites

Cited by 121 publications

References 38 publications

BDA-410: A novel synthetic calpain inhibitor active against blood stage malaria

BDA-410: A novel synthetic calpain inhibitor active against blood stage malaria

Overexpression of Leucyl Aminopeptidase in Plasmodium falciparum Parasites

Falcipain Cysteine Proteases Require Bipartite Motifs for Trafficking to the Plasmodium falciparum Food Vacuole

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