Isolation of merozoite rhoptries, identification of novel rhoptry-associated proteins from Plasmodium yoelii, P. chabaudi, P. berghei, and conserved interspecies reactivity of organelles and proteins with P. falciparum rhoptry-specific antibodies.
Rhoptries were isolated from merozoites of P. yoelii (17 XL), P. chabaudi adami and P. berghei (K-173), using sucrose gradient density centrifugation. Mouse antisera was prepared against the organelles and characterized. Antibodies specific for a known P. yoelii rhoptry protein were used to identify gradient fractions containing rhoptries and electron microscopy was used to confirm rhoptry enrichment and organelle morphology. Western blotting analysis of the gradients with organelle-specific antisera from each species, revealed several major cross-reactive interspecies protein bands of approximately 235, 210, 180, 160/170, 140, and 96-110 kDa, predominantly in densities of 1.12 and 1.15 g/ml. The parasite origin of the proteins was verified by immunoprecipitation, and reactive epitopes localized to the rhoptries by IEM. By Western blotting antisera specific for P. falciparum rhoptries reacted with protein bands of approximately 96-110 kDa in schizont extracts, and gradient fractions of density 1.12 and 1.15 g/ml from all three rodent malaria species, as well as with the rhoptries in P. yoelii, P. chabaudi, and P. berghei merozoites by IEM. We conclude that the three rodent malaria species and P. falciparum share conserved interspecies epitopes.
Department of Biology, Cleveland State University, Ohio 44115, USA. yellowe@biology.csuohio.edu
T Y
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Sam-Yellowe
R A
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Del Rio
H
H
Fujioka
M
M
Aikawa
J C
JC
Yang
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Yakubu
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000913:Antibodies, Protozoan / Q000276:immunology; D000953:Antigens, Protozoan / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D015153:Blotting, Western; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis / Q000737:chemistry / Q000276:immunology; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D007106:Immune Sera / Q000276:immunology; D008297:Male; D051379:Mice; D016253:Microscopy, Immunoelectron; D008970:Molecular Weight; D015388:Organelles / Q000737:chemistry / Q000276:immunology / Q000648:ultrastructure; D010962:Plasmodium berghei / Q000737:chemistry* / Q000276:immunology / Q000648:ultrastructure; D016788:Plasmodium chabaudi / Q000737:chemistry* / Q000276:immunology / Q000648:ultrastructure; D010967:Plasmodium yoelii / Q000737:chemistry* / Q000276:immunology / Q000648:ultrastructure; D011233:Precipitin Tests; D015800:Protozoan Proteins / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D016622:Silver Staining
D000911:Antibodies, Monoclonal; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D000939:Epitopes; D007106:Immune Sera; D015800:Protozoan Proteins; C065352:rhoptry associated protein, Plasmodium