Identification and Characterization of a Novel Plasmodium falciparum Merozoite Apical Protein Involved in Erythrocyte Binding and Invasion

Abstract

Proteins that coat Plasmodium falciparum merozoite surface and those secreted from its apical secretory organelles are considered promising candidates for the vaccine against malaria. In the present study, we have identified an asparagine rich parasite protein (PfAARP; Gene ID PFD1105w), that harbors a predicted signal sequence, a C-terminal transmembrane region and whose transcription and translation patterns are similar to some well characterized merozoite surface/apical proteins. PfAARP was localized to the apical end of the merozoites by GFP-targeting approach using an inducible, schizont-stage expression system, by immunofluorescence assays using anti-PfAARP antibodies. Immuno-electron microsopic studies showed that PfAARP is localized in the apical ends of the rhoptries in the merozoites. RBC binding assays with PfAARP expressed on COS cells surface showed that it binds to RBCs through its N-terminal region with a receptor on the RBC surface that is sensitive to trypsin and neuraminidase treatments. Sequencing of PfAARP from different P. falciparum strains as well as field isolates showed that the N-terminal region is highly conserved. Recombinant protein corresponding to the N-terminal region of PfAARP (PfAARP-N) was produced in its functional form in E. coli. PfAARP-N showed reactivity with immune sera from individuals residing in P. falciparum endemic area. The anti-PfAARP-N rabbit antibodies significantly inhibited parasite invasion in vitro. Our data on localization, functional assays and invasion inhibition, suggest a role of PfAARP in erythrocyte binding and invasion by the merozoite.

Plasmodium falciparumMerozoite Surface Protein 1 (MSP-1)-MSP-3 Chimeric Protein: Immunogenicity Determined with Human-Compatible Adjuvants and Induction of Protective Immune Response

“…In the absence of any direct correlates of protection against malaria, in vitro assays such as invasion inhibition and ADCI assays are used to assess the antiparasitic efficacy of the immune response generated to an immunogen (11,36,39,40,46). We found that purified Abs from sera of rabbits immunized with MSP-Fu 24 strongly inhibited the parasite invasion of not only homologous P. falciparum strain 3D7 but also strain FCR.…”

“…Antibodies to the recombinant proteins were tested for their reactivity with native parasite proteins by an immunofluorescence assay (IFA). The assay was performed essentially as described previously (36,46). Briefly, multispot parasite slides were made from P. falciparum (3D7) cultures, air dried, fixed with a mixture of acetone-methanol (9:1), and then blocked in blocking buffer (10% fetal calf serum [FCS] in PBS) for 1 h at 37°C.…”

“…Invasion inhibition assays were performed as described previously (36,46). Briefly, ring-stage malaria parasites were synchronized with sorbitol lysis and allowed to mature through to the schizont stage.…”

Host ICAMs play a role in cell invasion by Mycobacterium tuberculosis and Plasmodium falciparum

“…The effect of M5 on parasite invasion was evaluated on four different strains of P. falciparum (3D7, Dd2, HB3 and MCamp; see Supplementary Table 1). Invasion inhibition assay was performed as described previously 64 . Briefly, the haematocrit and parasitaemia of synchronized schizont stage cultures was adjusted to 2% and 1%, respectively.…”

Structural Insights into the Inactive Subunit of the Apicoplast-localized Caseinolytic Protease Complex of Plasmodium falciparum

“…Cryo-immunoelectron Microscopy-Immunoelectron microscopy was carried out on transgenic P. falciparum parasites expressing PfClpR-GFP at trophozoite stages as described earlier (28,37). Parasites were fixed in 4% paraformaldehyde, 0.04% glutaraldehyde in 1ϫ PBS at 4°C for 1 h and, subsequently, embedded in gelatin and infiltrated with a cryo-preservative and plasticizer (2.3 M sucrose, 20% polyvinyl pyrrolidone).…”