The late blood stages of the human malaria parasite, Plasmodium falciparum, carry a major surface antigen, p190, of molecular weight (Mr) 190,000. This antigenically variable protein is actively processed, first as the parasite matures and again when it is released into the blood stream and invades a new erythrocyte to initiate a cycle of growth. It elicits a strong immune response in man; all tested adult sera from endemic areas have antibodies against this protein. Our evidence indicates that purified p190 can alter the course of parasitaemia in monkeys with falciparum malaria. We have also succeeded in cloning part of the gene for p190 and expressing it in Escherichia coli. To this end we have developed a new technique, antibody select, which greatly simplifies final identification of expressing clones.
Saimiri monkeys immunized with a Plasmodium falciparum merozoite polypeptide of 41 kD mol wt are resistant to a blood challenge infection that induces a fulminant infection in control monkeys. The sera of the immunized monkeys reacted, as shown by the indirect immunofluorescence technique, with the apical part of the merozoites from five isolates or clones of P. falciparum. Whether the immunogen was dissolved in nonionic detergent (NP-40) or in sodium dodecyl sulfate (SDS) had a marked influence on the level of protection in immunized monkeys. Thus, monkeys immunized with the antigen solubilized in a nonionic detergent developed much lower parasitemia than monkeys immunized with denatured antigen (antigen eluted from SDS polyacrylamide gel electrophoresis).
Polypeptides expressed on the surface of merozoites, the invasive stage of the asexual blood cycle, are good candidates for the development of malaria vaccines. Five synthetic peptides with predetermined specificity deduced from a genomic DNA clone coding for the NH2-terminal portion of the main merozoite surface polypeptide of Plasmodium falciparum were evaluated for their capability to raise antibodies that react with the P. falciparum merozoites. Antibodies induced by two of the peptides (3 and 5) reacted with the membrane surfaces of seven of seven isolates of P. falciparum from different geographic areas. Antibodies against peptide 4, which contains a repeated amino acid sequence (Gly-Gly-Ser and Val-Ala-Ser), reacted with six of seven isolates. Structural analysis of the deduced polypeptides suggests that peptide 3 is exposed at the surface of merozoites. When it was used to immunize monkeys, three of the four animals were partially protected from a challenge infection that induced a fulminant infection in control animals.There is evidence to suggest that parasite components expressed at the surface of the invasive stages (sporozoite and merozoite) of Plasmodium falciparum, the most lethal malaria parasite species for humans, may be used for the development of malaria vaccines (1-5). Such vaccines require large amounts of antigens, which cannot be easily purified from the parasites but can be produced in large quantities by use of recombinant DNA technology (6-9).Previously, we have isolated a plasmid (pMC31-1) that codes for a portion of the schizont (180-200 kDa) and merozoite (83 kDa)-specific polypeptide from a cDNA library constructed from mRNA purified from the asexual blood stages of P. falciparum (isolate SGE2 from Zaire). The 83-kDa protein is a processed product of the 180-to 200-kDa polypeptide and it is expressed at the surface of the merozoites (5, 9). The potential value for vaccine development of the malaria-specific protein encoded by pMC31-1 plasmid is substantiated by the finding that antisera raised against lysates of pMC31-1-containing bacteria react with the merozoite surface of five out of five P. falciparum isolates from various geographic locations and by immunization trials showing that monkeys immunized with the 180-to 200-kDa polypeptide are resistant to a blood-induced challenge infection (10). The aim of this study was to evaluate the possibility of raising antibodies against this protein by immunization with synthetic peptides of predetermined specificity containing either unique or repeated amino acid sequences deduced from the cDNA sequence. Antisera raised against the synthetic peptides were tested for their capacity to react with asexual blood stage parasites and with the 180-to 200-kDa polypeptide and its processed products. Finally, a synthetic polypeptide was selected for an immunization experiment with monkeys.
MATERIALS AND METHODSParasites. Asynchronous in vitro cultures of seven P. falciparum isolates-SGE2 (from Zaire), FUP Palo Alto (Uganda), FCR3 clone A2 ...
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