Purification and characterization of 37-kilodalton proteases from Plasmodium falciparum and Plasmodium berghei which cleave erythrocyte cytoskeletal components.
Cytosoluble 100,000 X g extracts from Plasmodium berghei or Plasmodium falciparum infected red blood cells were shown to hydrolyze erythrocyte spectrin. By Fast Protein Liquid Chromatography (FPLC), these enzymes were purified and exhibited a pI of 4.5 and Mr of 37,000 using SDS-PAGE under reducing conditions. An immunochemical enzyme assay using anti-spectrin antibodies was developed. The optimal activity using spectrin as substrate was at pH 5.0, and the enzymes were strongly inhibited by HgCl2, ZnCl2, chymostatin, leupeptin and aprotinin, and moderately by pepstatin. These properties of the Pf37 and Pb37 proteases differ from the Plasmodium lophurae and P. falciparum 'cathepsin D-like' enzymes and from the serine or cysteine neutral proteases previously described in P. falciparum and P. berghei infected red blood cells. While the Pf37 and Pb37 enzymes cleaved spectrin preferentially, degradation of band 4.1 was also observed with high concentration of enzyme. The parasite origin of the Pf37 protease was clearly demonstrated, since purified radiolabeled enzyme was active on spectrin. A high-molecular-weight polymer (greater than 240 kDa) was often observed on incubating purified spectrin and Pf37 protease. The breakdown of erythrocyte cytoskeletal components could be of interest in the release of merozoites from segmented schizonts or during the process of invasion of erythrocytes by merozoites.
Deguercy
A
A
Laboratoire de Biologie Cellulaire, URA CNRS 80, Université de Poitiers, France.
Hommel
M
M
Schrével
J
J
D000818:Animals; D004912:Erythrocytes / Q000378:metabolism; D007124:Immunoenzyme Techniques; D008715:Methionine / Q000378:metabolism; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000302:isolation & purification*; D010962:Plasmodium berghei / Q000201:enzymology*; D010963:Plasmodium falciparum / Q000201:enzymology*; D013049:Spectrin / Q000378:metabolism*; D013462:Sulfur Radioisotopes
D013462:Sulfur Radioisotopes; D013049:Spectrin; D008715:Methionine; D010447:Peptide Hydrolases