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Sequence analysis of the Rhop-3 gene of Plasmodium berghei and P. chabaudi, reactivity of Rhop-3 protein within isolated rhoptries and binding of Rhop-3 to mouse erythrocytes

Sequence analysis of the Rhop-3 gene of Plasmodium berghei and P. chabaudi, reactivity of Rhop-3 protein within isolated rhoptries and binding of Rhop-3 to mouse erythrocytes

Journal of Protozoology Research 10(2): 71-89

The 110 kDa Rhop-3 rhoptry protein of Plasmodium falciparum is secreted into the erythrocyte membrane during invasion. It is an erythrocyte binding protein that is non-covalently associated with two other proteins, the 140 kDa Rhop-1 and the 130 kDa Rhop-2. We identified the Rhop-3 gene homologue in P. yoelii and demonstrated that the C-terminus is highly conserved. In order to identify the Rhop-3 gene homologue in P. berghei and P. chabaudi, a set of primers were designed based on the cDNA sequence of clone Y1412 of P. yoelii and used to amplify genomic DNA from P. berghei and P. chabaudi by polymerase chain reaction (PCR). Analysis of the DNA and deduced amino acid sequence demonstrated sequence homology to P. falciparum Rhop-3. We examined the distribution of Rhop-3 epitopes within isolated rhoptries of the three rodent Plasmodium species and P. falciparum, and investigated the erythrocyte binding property of rodent Plasmodium Rhop-3. We also evaluated the immunogenicity of isolated rhoptries from the rodent Plasmodium species following treatment of the organelles with 100 mM sodium carbonate, pH 11.5 and 0.5% SDS, to stimulate antibodies to proteins not accessible when untreated organelles are used for immunization. We show that the integrity of the isolated organelles was stable and the organelles were reactive with Rhop-3 specific antisera. In addition, the rodent Plasmodium Rhop-3 protein bound to mouse erythrocytes and was recognized by Rhop-3 specific antibody. Furthermore, treated organelles stimulated antisera that recognized additional rhoptry proteins not seen when antisera prepared against whole untreated organelles were examined by western blotting. Taken together, these data suggest that malaria vaccine studies using the Rhop-3 protein can be performed directly in vivo using the rodent Plasmodium models.

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Accession: 003930821

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