MAEBL Plasmodium falciparum protein peptides bind specifically to erythrocytes and inhibit in vitro merozoite invasion

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Abstract

MAEBL is an erythrocyte binding protein located in the rhoptries and on the surface of mature merozoites, being expressed at the beginning of schizogony. The structure of MAEBL originally isolated from rodent malaria parasites suggested a molecule likely to be involved in invasion. We thus became interested in identifying possible MAEBL functional regions. Synthetic peptides spanning the MAEBL sequence were tested in erythrocyte binding assays to identify such possible MAEBL functional regions. Nine high activity binding peptides (HABPs) were identified: two were found in the M1 domain, one was found between the M1 and M2 regions, five in the erythrocyte binding domain (M2), and one in the protein’s repeat region. The results showed that peptide binding was saturable; some HABPs inhibited in vitro merozoite invasion and specifically bound to a 33 kDa protein on red blood cell membrane. HABPs’ possible function in merozoite invasion of erythrocytes is also discussed.

Section snippets

Materials and methods

Peptide synthesis. One hundred and three sequential 20-mer peptides, corresponding to the complete P. falciparum MAEBL protein amino acid sequence [GenBank Accession No. NP_701342], were synthesised by the solid phase multiple peptide system [16], [17]; t-Boc amino acids (Bachem); and MBHA resin (0.7 meq/g) were used. Peptides were cleaved by the low-high HF technique [18], purified by RP-HPLC, lyophilised, and analysed by MALDI-TOF mass spectrometry. Tyrosine was added to those peptides which

MAEBL peptides bind specifically to human erythrocytes

Binding assays were used to determine specific erythrocyte binding activity for 103 synthetic peptides covering the total length of the MAEBL protein [GenBank Accession No. NP_701342]. Peptide binding activity was defined as being the amount (pmol) of peptide that bound specifically to erythrocyte per added peptide (pmol). High activity binding peptides (HABPs) were defined as those peptides showing activity greater than or equal to 2%, according to previously established criteria [20], [21],

Discussion

A better understanding of the complex process of P. falciparum merozoite invasion requires that the numerous potential parasite ligands become identified and characterised. Our studies have required an innovative methodology to be employed in the search for sequences which can be precisely modified for use as candidates for a vaccine against malaria [28], [29], [30], [31]. Some of the important ligand determinants for invasion belong to the members of the ebl family [11]. Differences in

Acknowledgements

This research project was supported by the President of the Republic of Colombia’s office and the Colombian Ministry of Public Health. We thank Jason Garry for reading the manuscript.

References (40)

  • S.A Dolan et al.

    Glycophorin B as an EBA-175 independent Plasmodium falciparum receptor on human erythrocytes

    Mol. Biochem. Parasitol.

    (1994)
  • G.H Mitchell et al.

    Invasion of erythrocytes by Plasmodium falciparum malaria parasites: evidence for receptor heterogeneity and two receptors

    Blood

    (1986)
  • M.E Perkins et al.

    Erythrocyte receptor recognition varies in Plasmodium falciparum isolates

    Mol. Biochem. Parasitol.

    (1988)
  • C.E Chitnis

    Molecular insights into receptors used by malaria parasites for erythrocyte invasion

    Curr. Opin. Hematol.

    (2001)
  • J.A Dvorak et al.

    Invasion of erythrocytes by malaria merozoites

    Science

    (1975)
  • M Aikawa et al.

    Erythrocyte entry by malarial parasites. A moving junction between erythrocyte and parasite

    J. Cell Biol.

    (1978)
  • G.E Ward et al.

    The invasion of erythrocytes by malarial merozoites

  • L.H Bannister et al.

    Lamellar membranes associated with rhoptries in erythrocytic merozoites of Plasmodium knowlesi: a clue to the mechanism of invasion

    Parasitology

    (1986)
  • J.H Adams et al.

    A family of erythrocyte binding proteins of malaria parasites

    Proc. Natl. Acad. Sci. USA

    (1992)
  • S.H Kappe et al.

    A family of chimeric erythrocyte binding proteins of malaria parasites

    Proc. Natl. Acad. Sci. USA

    (1998)
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