Neutral proteases involved in the reinvasion of erythrocytes by Plasmodium merozoites.
Neutral proteases of Plasmodium sp erythrocytic stages were studied by means of a sensitive fluorogenic method and gelatin-SDS-PAGE. The substrates gluconoyl-Val-Leu-Gly-Lys(or Arg)-3-amido-9-ethylcarbazole were selectively hydrolyzed by an endopeptidase from rodent Plasmodium berghei (Pb) and Plasmodium chabaudi (Pc) and from human Plasmodium falciparum (Pf) parasites. These endopeptidases were purified from 100,000-g soluble schizont extract by high pressure liquid chromatography; they have a similar Mr of 68,000 in SDS-PAGE, and an optimal activity at pH 7.4. The Pb 68 and Pf 68 endopeptidases were localized in schizonts and also in merozoites as shown by indirect immunofluorescence on Pb merozoites and by the identification of the Pf 68 endopeptidase activity in free viable merozoites. The Pb 68 and Pf 68 endopeptidases belong to the class of cysteine proteases. Analysis by gelatin-SDS-PAGE of a Pb 68 endopeptidase-enriched fraction showed a reproducible 95,000 proteolytic band. The initial extracts showed a similar 95,000 proteolytic band, and also 2 other 90,000 and 85,000 major bands. During reinvasion experiments, it was possible to recover a 95,000 and a 40,000 protease band from supernates of cultures grown in a semidefined medium without serum. Hydrophilic peptide derivatives related to the substrate of Pf 68 endopeptidase are shown to be potential inhibitors of the Pf reinvasion process in vitro.
Schrevel
J
J
Laboratoire de Biologie Cellulaire, URA CNRS no. 80, Université de Poitiers, France.
Grellier
P
P
Mayer
R
R
Monsigny
M
M
D000818:Animals; D000953:Antigens, Protozoan / Q000276:immunology; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000469:parasitology*; D010447:Peptide Hydrolases / Q000032:analysis*; D010961:Plasmodium / Q000201:enzymology* / Q000254:growth & development; D011480:Protease Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity
D000953:Antigens, Protozoan; D003470:Culture Media; D011480:Protease Inhibitors; D010447:Peptide Hydrolases