Purification of Plasmodium lophurae cathepsin D and its effects on erythrocyte membrane proteins

Sherman, Irwin W.; Tanigoshi, Linda

doi:10.1016/0166-6851(83)90044-0

Cited by 41 publications

(16 citation statements)

References 21 publications

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“…This may merely be an artifact of isolation or fixation. Alternatively, considering the paramagnetic properties of the iron in hemozoin (44), a reasonable hypothesis is that the pH gradient (24,25). We would suggest that a likely explanation is that the TCA solubility assay used in these studies is insensitive to endoproteolytic clipping so that, as the proteases that generate smaller fragments were purified away, activity in their assays disappeared.…”

Section: Discussionmentioning

confidence: 99%

Hemoglobin degradation in the malaria parasite Plasmodium falciparum: an ordered process in a unique organelle.

Goldberg

Slater

Cerami

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

418

272

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The malaria parasite Plasmodium falkiparum uses host erythrocyte hemoglobin as a major nutrient source. We report the purification of P. falctprum digestive vacuoles and characterization of the degradative process therein. Vacuoles were isolated by a combination of differential centrifugation and density gradient separation. The pure vacuoles were capable of degrading hemoglobin to small fragments with a pH optimum of 5-5.5. Proteolysis in the vacuoles appears to be an ordered process, requiring an aspartic protease to clip intact hemoglobin before other proteolytic activities can function efficiently. The vacuoles do not contain other hydrolases commonly found in lysosomes and therefore appear to be unique proteolytic organelles designed specifically to degrade hemoglobin.

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Section: Discussionmentioning

confidence: 99%

Hemoglobin degradation in the malaria parasite Plasmodium falciparum: an ordered process in a unique organelle.

Goldberg

Slater

Cerami

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

418

272

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“…This process involves the transport of erythrocyte cytoplasm to acidic food vacuoles, where the heme component of hemoglobin is processed into hemozoin, and globin is hydrolized to free amino acids (2). As is the case with mammalian lysosomes, malarial food vacuoles appear to contain both cysteine (3) and aspartic (4)(5)(6) proteinases, both of which may participate in the hydrolysis of globin (7).…”

Section: Introductionmentioning

confidence: 99%

Inhibition of a Plasmodium vinckei cysteine proteinase cures murine malaria.

Rosenthal¹,

Lee²,

Smith³

1993

J. Clin. Invest.

155

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Intraerythrocytic malaria parasites degrade hemoglobin as a principal source of amino acids for parasite protein synthesis. We have previously identified a Plasmodium falciparum trophozoite cysteine proteinase as a putative hemoglobinase and shown that specific inhibitors of this proteinase block the hydrolysis of globin and the development of cultured parasites. We now show that the murine malaria parasite Plasmodium vinckei has an analogous cysteine proteinase with similar biochemical properties to the P. falciparum proteinase, including an acid pH optimum, a preference for the peptide proteolytic substrate benzyloxycarbonyl (Z)-Phe-Arg-7-amino4-methylcoumarin, and nanomolar inhibition by seven peptide fluoromethyl ketone proteinase inhibitors. Thus, P. vinckei offers a model system for the in vivo testing of the antimalarial properties of cysteine proteinase inhibitors. One of the proteinase inhibitors studied, morpholine urea (Mu)-Phe-Homophenylalanine (HPhe)-CH2F strongly inhibited the P. vinckei cysteine proteinase in vitro and rapidly blocked parasite cysteine proteinase activity in vivo. When administered four times a day for 4 d to P. vinckei-infected mice, Mu-Phe-HPhe-CH2F elicited long-term cures in 80% of the treated animals. These results show that peptide proteinase inhibitors can be effective antimalarial compounds in vivo and suggest that the P. falciparum cysteine proteinase is a promising target for chemotherapy. (J.

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“…[10][11][12][13][14][15] Proteolytic breakdown of these proteins results in red cell lysis and thus could potentially provide a mechanism for merozoite release from RBCs. In this regard, we have recently shown that P falciparum-derived cysteine protease falcipain-2 (FP-2) cleaves host erythrocyte membrane skeletal proteins at neutral pH.…”

Section: Introductionmentioning

confidence: 99%

Plasmodium falciparum cysteine protease falcipain-2 cleaves erythrocyte membrane skeletal proteins at late stages of parasite development

Hanspal

Dua

Takakuwa

et al. 2002

Blood

108

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Plasmodium falciparum-derived cysteine protease falcipain-2 cleaves host erythrocyte hemoglobin at acidic pH and specific components of the membrane skeleton at neutral pH. Analysis of stage-specific expression of these 2 proteolytic activities of falcipain-2 shows that hemoglobinhydrolyzing activity is maximum in early trophozoites and declines rapidly at late stages, whereas the membrane skeletal protein hydrolyzing activity is markedly increased at the late trophozoite and schizont stages. Among the erythrocyte membrane skeletal proteins, ankyrin and protein 4.1 are cleaved by native and recombinant falcipain-2 near their C-termini. To identify the precise peptide sequence at the hydrolysis site of protein 4.1, we used a recombinant construct of protein 4.1 as substrate followed by MALDI-MS analysis of the cleaved product. We show that falcipain-2-mediated cleavage of protein 4.1 occurs immediately after lysine 437, which lies within a region of the spectrinactin-binding domain critical for erythrocyte membrane stability. A 16-mer peptide containing the cleavage site completely inhibited the enzyme activity and blocked falcipain-2-induced fragmentation of erythrocyte ghosts. Based on these results, we propose that falcipain-2 cleaves hemoglobin in the acidic food vacuole at the early trophozoite stage, whereas it cleaves specific components of the red cell skeleton at the late trophozoite and schizont stages. It is the proteolysis of skeletal proteins that causes membrane instability, which, in turn, facilitates parasite release in vivo. IntroductionPlasmodium falciparum causes the most severe form of human malaria and is becoming increasingly resistant to available antimalarial drugs. New chemotherapy-based approaches to fight the disease are therefore urgently needed. Parasite proteases that are involved in P falciparum development appear to be good targets. Mounting evidence suggests that cysteine proteases are involved in host cell rupture and release of merozoites. In the presence of such inhibitors, merozoites mature normally but are unable to escape from host erythrocytes. [1][2][3][4] The cluster of merozoites inside a red blood cell (RBC) is enclosed within 2 membranes: an inner parasitophorous vacuole membrane (PVM) and an outer RBC membrane. The rupture of these 2 membranes apparently releases the merozoites for another round of RBC invasion. In a recent study by Salmon et al,5 the authors propose a 2-step process for parasite release from the host erythrocyte: an initial exit of merozoites enclosed within the PVM followed by a rapid escape from the PVM by a proteolysisdependent mechanism. This study suggests that the RBC membrane is lost independently of the PVM. In another report, Winograd et al 6 used videomicroscopy to study the release of merozoites and concluded that an aperture is made through the PVM and RBC membranes to allow merozoites to exit in an orderly fashion. Merozoites were released together with the residual body containing hemozoin, leaving behind the red cell membrane and some...

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Purification of Plasmodium lophurae cathepsin D and its effects on erythrocyte membrane proteins

Cited by 41 publications

References 21 publications

Hemoglobin degradation in the malaria parasite Plasmodium falciparum: an ordered process in a unique organelle.

Hemoglobin degradation in the malaria parasite Plasmodium falciparum: an ordered process in a unique organelle.

Inhibition of a Plasmodium vinckei cysteine proteinase cures murine malaria.

Plasmodium falciparum cysteine protease falcipain-2 cleaves erythrocyte membrane skeletal proteins at late stages of parasite development

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