Lysis of Plasmodium falciparum by ferriprotoporphyrin IX and a chloroquine-ferriprotoporphyrin IX complex

Fitch, Coy D.; Chevli, Rekha; Banyal, H. S.; Phillips, Gary; Pfaller, Michael A.; Krogstad, Donald J.

doi:10.1128/aac.21.5.819

Cited by 147 publications

(62 citation statements)

References 18 publications

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“…The cells were pelleted down and the parasites, essentially at the trophozoite stage, were isolated by treatment with 0.15% (w/v) saponin (9) or osmotic lysis (10) with slight modifications. For the latter procedure, packed infected erythrocytes were treated with 40 volumes of cold 10 mM potassium phosphate buffer, pH 7.4, mixed thoroughly and allowed to stand on ice for 10 min.…”

Section: Methodsmentioning

confidence: 99%

δ-Aminolevulinic Acid Dehydratase from Plasmodium falciparum

Dhanasekaran

Chandra

Sagar

et al. 2004

Journal of Biological Chemistry

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The heme biosynthetic pathway of the malaria parasite is a drug target and the import of host ␦-aminolevulinate dehydratase (ALAD), the second enzyme of the pathway, from the red cell cytoplasm by the intra erythrocytic malaria parasite has been demonstrated earlier in this laboratory. In this study, ALAD encoded by the Plasmodium falciparum genome (PfALAD) has been cloned, the protein overexpressed in Escherichia coli, and then characterized. The mature recombinant enzyme (rPfALAD) is enzymatically active and behaves as an octamer with a subunit M r of 46,000. The enzyme has an alkaline pH optimum of 8.0 to 9.0. rPfALAD does not require any metal ion for activity, although it is stimulated by 20 -30% upon addition of Mg 2؉ . The enzyme is inhibited by Zn 2؉ and succinylacetone. The presence of PfALAD in P. falciparum can be demonstrated by Western blot analysis and immunoelectron microscopy. The enzyme has been localized to the apicoplast of the malaria parasite. Homology modeling studies reveal that PfALAD is very similar to the enzyme species from Pseudomonas aeruginosa, but manifests features that are unique and different from plant ALADs as well as from those of the bacterium. It is concluded that PfALAD, while resembling plant ALADs in terms of its alkaline pH optimum and apicoplast localization, differs in its Mg 2؉ independence for catalytic activity or octamer stabilization. Expression levels of PfALAD in P. falciparum, based on Western blot analysis, immunoelectron microscopy, and EDTA-resistant enzyme activity assay reveals that it may account for about 10% of the total ALAD activity in the parasite, the rest being accounted for by the host enzyme imported by the parasite. It is proposed that the role of PfALAD may be confined to heme synthesis in the apicoplast that may not account for the total de novo heme biosynthesis in the parasite.Studies in this laboratory had demonstrated that the malaria parasites (Plasmodium falciparum and Plasmodium berghei) are capable of heme biosynthesis de novo, despite acquiring large amounts of heme from the host red cell hemoglobin in the intraerythrocytic stage (1, 2). Inhibition of the de novo heme biosynthetic pathway leads to death of the parasite and the pathway is therefore a drug target (1-3). Studies on the enzymes of the heme-biosynthetic pathway have revealed that the first enzyme, ␦-aminolevulinate synthase, is coded for by the parasite genome and is localized in the parasite mitochondrion (4, 5). Studies from this laboratory have shown that the second enzyme of the pathway, ␦-aminolevulinate dehydratase (ALAD), 1 in the parasite is of host origin and evidence was provided to show that the enzyme is translocated (imported) from the host red cell into the parasite and is functional (2, 3).At the same time genome sequence data available for P. falciparum indicates that the parasite genome codes for ALAD (PfALAD) and other enzymes of the heme-biosynthetic pathway. Sato et al. (6) have shown by phylogenetic amino acid sequence analysis derived from truncated...

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Section: Methodsmentioning

confidence: 99%

δ-Aminolevulinic Acid Dehydratase from Plasmodium falciparum

Dhanasekaran

Chandra

Sagar

et al. 2004

Journal of Biological Chemistry

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“…The biochemical consequences of nonsequestered heme accumulation are profound and include membrane lysis (18) and protease inhibition (19,20). It was recently recognized that one defense plasmodia have developed against these drugs is efficient drug excretion out of the digestive vacuole (21); however, there may be other adaptations in the chloroquine-resistant strains.…”

Section: X-ray Diffraction Of Hemozoin and ␤-Hematin 714mentioning

confidence: 99%

Characterization of the Products of the Heme Detoxification Pathway in Malarial Late Trophozoites by X-ray Diffraction

Bohle

Dinnebiér

Madsen

et al. 1997

Journal of Biological Chemistry

151

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In a process inhibited by the quinoline antimalarial drugs, Plasmodia detoxify heme released during the degradation of hemoglobin by aggregating it into malarial pigment, an insoluble crystalline heme coordination polymer. Synchrotron x-ray powder diffraction patterns for intact desiccated malarial trophozoites and synthetic ␤-hematin have been measured; both materials correspond to a single crystalline triclinic lattice with unit cell parameters a ‫؍‬ 12.2176(4), b ‫؍‬ 14.7184(5), c ‫؍‬ 8.0456(3) Å; ␣ ‫؍‬ 90.200(2), ␤ ‫؍‬ 96.806(3), ␥ ‫؍‬ 97.818(3)°a nd Z ‫؍‬ 2. These results unambiguously demonstrate that hemozoin crystallites are identical to synthetic ␤-hematin.Heme is a potent multifunction regulator whose biochemical levels and distribution are precisely controlled on both intraand extracellular levels (1). Efficient regulation of heme is particularly critical for intra-erythrocytic parasites such as plasmodia which process large quantities of heme in the postinvasion digestion of the erythrocyte's hemoglobin. Plasmodia, which lack heme oxygenases, detoxify heme by sequestering it into an insoluble heme aggregate termed malarial pigment or hemozoin (2). The quinoline-based family of antimalarials interfere with this process by an as yet unknown mechanism that has recently come under intense scrutiny as part of the effort to combat the spread of chloroquine-resistant strains of Plasmodium. A variety of spectroscopic and bioanalytical techniques indicate that hemozoin is similar to the synthetic aggregated heme phase ␤-hematin, which is thought to form strands of hemes linked by propionate oxygen-iron bonds as well as interstrand propionate hydrogen bonds, Fig. 1 (3-5). Characterization of the carboxylate stretching bands for the propionic acid side chains by IR and Raman spectroscopy provides the best evidence for the presence of iron-oxygen bonds to the propionate side chains (3-5). Unfortunately, crystallographic characterization of these heme aggregates has been hampered by the phase heterogeneity of many synthetic preparations as well as by the small size of the synthetic and natural crystallites isolated from either trophozoites and infected hosts (6). High resolution powder diffraction has been used extensively for the solution of many structural problems (7), and it can solve problems posed by diffraction from microcrystalline phases. In this communication we describe the characterization of ␤-hematin derived from both synthetic and natural sources and provide new unambiguous evidence that the heme aggregate present in late stage trophozoites is ␤-hematin. EXPERIMENTAL PROCEDURESCirca 3 ϫ 10 9 chloroquine-susceptible Plasmodium falciparum late trophozoites of the 3D7 clone (NF54 strain) were synchronized with sorbitol and allowed to achieve a 92% level of parasitemia (8). The resulting intact trophozites were isolated by rapid cooling in liquid nitrogen, and the frozen suspension of cells was lyophilized by freezedrying in vacuo at Ϫ10°C. In general this freeze-drying process required 12-14 h and res...

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“…Hemin can cause lysis of mouse [1,2] and human erythrocytes [3] and of malaria parasites [4,5]. Other types of cells were demonstrably resistant to this lytic effect (see Ref.…”

Section: Introductionmentioning

confidence: 99%

“…The combined toxic effect of heroin and the antimalarial drug chloroquine on cell membranes has been suggested to underly the pharmacological mode of action of this drug [16]. Hemin forms complexes with antimalarial drugs both in aqueous [1,17] and apolar [18] phases and in the presence of these complexes erythrocytes and malaria parasites lyse [1,4,5]. These observations, in conjunction with the present resurgence of malaria which is mostly due to chloroquine resistance and the lack of cross resistance to other 4-aminoquinolines [19,20], underscores the necessity for further probing of the molecular details of the interaction of hemin, drugs and heroin-drug complexes with phospholipids.…”

Section: Introductionmentioning

confidence: 99%

Interactions of hemin, antimalarial drugs and hemin-antimalarial complexes with phospholipid monolayers

Ginsburg

Demel

1984

Chemistry and Physics of Lipids

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Hemin, antimalarial drugs and complexes formed between them, have demonstrable effects on biological membranes. Using the phospholipid monolayer model, we show that hemin intercalates into the membrane and increases its surface pressure, depending on the lipid composition and the initial surface pressure: negative surface charges and particularly looser compaction of the phospholipids reduce the effect of hemin. With increasing surface pressure hemin tends to intercalate as a monomer, and the half-saturation concentration of its effect increases exponentially. The antimalarial monovalent drugs quinine and mefloquine, but not chloroquine, also penetrate into the membrane and expand it. All three drugs markedly increase the effect of hemin, but chloroquine reduces the effect in monolayers composed of unsaturated phospholipids. The drugs' effect is mostly due to an increase in the maximal surface pressure and suggests a complexation of heroin and drug within the membrane phase. Preformed hemindrug complexes decrease the half-saturation concentration of the effect and suggest that the complexes adsorb to the membrane, releasing the hemin through an apolar continuum into the phospholipid phase. The implications of the results to the membrane toxicity mechanism proposed for the molecular mode of action of antimalarial drugs are discussed.

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Lysis of Plasmodium falciparum by ferriprotoporphyrin IX and a chloroquine-ferriprotoporphyrin IX complex

Cited by 147 publications

References 18 publications

δ-Aminolevulinic Acid Dehydratase from Plasmodium falciparum

δ-Aminolevulinic Acid Dehydratase from Plasmodium falciparum

Characterization of the Products of the Heme Detoxification Pathway in Malarial Late Trophozoites by X-ray Diffraction

Interactions of hemin, antimalarial drugs and hemin-antimalarial complexes with phospholipid monolayers

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