Baculovirus-mediated expression of Plasmodium falciparum erythrocyte binding antigen 175 polypeptides and their recognition by human antibodies.
The erythrocyte binding antigen EBA-175 is a 175-kDa Plasmodium falciparum protein which mediates merozoite invasion of erythrocytes in a sialic acid-dependent manner. The purpose of this study was to produce recombinant EBA-175 polypeptide domains which have previously been identified as being involved in the interaction of EBA-175 with erythrocytes and to determine whether these polypeptides are recognized by malaria-specific antibodies. The eba-175 gene was cloned by PCR from genomic DNA isolated from the 3D7 strain of P. falciparum. The predicted protein sequence was highly conserved with that predicted from the published eba-175 gene sequences from the Camp and FCR-3 strains of P. falciparum and contained the F segment divergent region. Purified recombinant EBA-175 polypeptide fragments, expressed as glutathione S-transferase fusion proteins in insect cells by using the baculovirus system, were recognized by antibodies present in serum from a drug-cured, malaria-immune Aotus nancymai monkey. The fusion proteins were also recognized by antibodies present in sera from individuals residing in areas where malaria is endemic. In both cases the antibodies specifically recognized the EBA-175 polypeptide portion of the fusion proteins. Antibodies raised in rabbits immunized with the recombinant fusion proteins recognized parasite proteins present in schizont-infected erythrocytes. Our results suggest that these regions of the EBA-175 protein are targets for the immune response against malaria and support their further study as possible vaccine components.
Daugherty
J R
JR
Department of Immunology, Walter Reed Army Institute of Research, Washington, D.C. 20307-5100, USA.
Murphy
C I
CI
Doros-Richert
L A
LA
Barbosa
A
A
Kashala
L O
LO
Ballou
W R
WR
Snellings
N J
NJ
Ockenhouse
C F
CF
Lanar
D E
DE
D000595:Amino Acid Sequence; D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology*; D000953:Antigens, Protozoan / Q000276:immunology*; D001029:Aotus trivirgatus / Q000276:immunology; D016367:Baculoviridae; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology; D010963:Plasmodium falciparum / Q000235:genetics / Q000276:immunology*; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000276:immunology; D018411:Spodoptera; D013329:Structure-Activity Relationship
D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D002352:Carrier Proteins; D010455:Peptides; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C065668:erythrocyte-binding antigen 175, Plasmodium