A Plasmodium falciparum antigen containing clusters of asparagine residues (original) (raw)
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Nucleic Acids Research, 1986
We describe a cDNA clone derived from mRNA of asexual blood-stages of the malaria parasite Plasmodium falciparum. This clone, designated Ag319, expresses a P.falciparum antigen fused to 0-galactosidase in Escherichia coli. Human antibodies from Papua New Guinea were affinity-purified by adsorption to extracts of Ag319 immobilized on CNBr-Sepharose. The antibodies reacted predominantly with P.falciparum polypeptides of Mr 220,000 and 160,000, and a number of ill-defined lower molecular weight species. Antibodies reacted in indirect immunofluorescence with all asexual blood-stages although the antigen appeared to be most abundance in the schizont. Surprizingly the antibodies also reacted with sporozoites. The amino acid sequence predicted from the complete nucleotide sequence of this clone is remarkable because 40% of the residues are Asn, and so the antigen has been termed the Asparagine-Rich Protein (ARP). Like other P.falciparum antigens, ARP contains tandemly repetitive sequences, based on the tetrapeptide Asn-Asn-Asn-Met and we have confirmed that these represent natural epitopes by reaction of the corresponding synthetic peptides with human antibodies. Surprisingly, ARP is also rich in Asn outside the tandem repeats.
2009
The genetic mutation in malaria parasite strain causing diversity characteristic of each strain. Many of the antigens, that P. falciparum express during their life cycle, particularly the asexual blood-stages, are antigenically diverse. The two major causes of antigenic diversity are allelic polymorphisms and antigenic variation. Each stage has different antigens that lead to protective immunity and in many cases; these antigens are not expressed at other stages of the life cycle. The TS.Ag isolated from P. falciparum 2300 strain containing mature asexual stages of parasite. Characterization of this antigen by SDS-PAGE and western blott methods has been performed to find out a malaria vaccine candidate for Indonesia locally. Comparison of the protein content of TS.Ag with R.Ag (ring form stage antigen) and RTS.Ag (ring form, trophozoite and schizont antigen) showed that, 14.5 kDa protein present in these three antigens. Western blott analysis of these antigens showed that, only 14.5...
The EMBO Journal, 1984
*To whom reprint requests should be sent Communicated by K. Murray A cDNA library was constructed in pBR322 using mRNA from blood stages of a Papua New Guinean isolate of Plasmodium falciparum. Expression of parasite antigens was not directly detectable by conventional immunological assays. To circumvent this, mice were immunized with lysates of cDNA clones, and the antisera raised were assayed for anti-parasite reactivity. One cDNA clone was identified which reliably elicited antibodies to P. falciparum. The mouse antisera were used to characterize the native P. falciparum protein as a 120-kd protein, which is antigenic during natural infection. The protein occurs in late trophozoite and schizont stages and is found in isolates of the parasite from widely separated geographical areas. The genomic context of the antigen gene is conserved in the different isolates.
The delineation of putatively protective and immunogenic epitopes in vaccine candidate proteins constitutes a major research effort towards the development of an effective malaria vaccine. By virtue of its role in the formation of the immune clusters of merozoites, its location on the surface of merozoites, and its highly conserved nature both at the nucleotide sequence level and the amino acid sequence level, the antigen which contains repeats of acidic and basic residues (ABRA) of the human malaria parasite Plasmodium falciparum represents such an antigen. Based upon the predicted amino acid sequence of ABRA, we synthesized eight peptides, with six of these (AB-1 to AB-6) ranging from 12 to 18 residues covering the most hydrophilic regions of the protein, and two more peptides (AB-7 and AB-8) representing its repetitive sequences. We found that all eight constructs bound an appreciable amount of antibody in sera from a large proportion of P. falciparum malaria patients; two of these peptides (AB-1 and AB-3) also elicited a strong proliferation response in peripheral blood mononuclear cells from all 11 human subjects recovering from malaria. When used as carrierfree immunogens, six peptides induced a strong, boostable, immunoglobulin G-type antibody response in rabbits, indicating the presence of both B-cell determinants and T-helper-cell epitopes in these six constructs. These antibodies specifically cross-reacted with the parasite protein(s) in an immunoblot and in an immunofluorescence assay. In another immunoblot, rabbit antipeptide sera also recognized recombinant fragments of ABRA expressed in bacteria. More significantly, rabbit antibodies against two constructs (AB-1 and AB-5) inhibited the merozoite reinvasion of human erythrocytes in vitro up to ϳ90%. These results favor further studies so as to determine possible inclusion of these two constructs in a multicomponent subunit vaccine against asexual blood stages of P. falciparum.
Journal of Clinical Microbiology, 1987
Pf155 is a merozoite-derived polypeptide antigen which the parasite Plasmodium falciparum deposits in the membranes of erythrocytes at invasion. Eleven laboratory strains or clones of P. falciparum and a large number of isolates obtained from patients from different parts of the world were studied for antigenic diversity in Pf155. Immunoglobulin G antibodies from different serum samples from P. falciparum-infected donors were affinity purified on monolayers of glutaraldehyde-fixed and air-dried erythrocytes infected with P. falciparum of different origins and tested in different combinations by immunoblotting, reinvasion inhibition, and a modified immunofluorescence procedure in which the membranes of recently infected erythrocytes were stained. Similar experiments were performed with monoclonal and oligoclonal antibodies specific for different epitopes in the C-terminal region of Pf155. No strain- or isolate-associated antigenic diversity or size variation of Pf155 was detected, in...
Infection and immunity, 1994
A differential serological screen of a lambda gt11 cDNA expression library of Plasmodium falciparum was performed in an attempt to identify novel and putative host-protective antigens of the parasite. The screening was done with two categories of sera: (i) acute-phase sera obtained from smear-positive acutely infected P. falciparum patients from various regions in India and (ii) immune sera taken from healthy, permanent adult residents of P. falciparum-endemic rural districts of Orissa in eastern India. These adults had not suffered from any clinical malarial symptoms for at least the previous 3 years at the time of serum collection. Sixty-five clones obtained by screening the lambda gt11 library with two immune serum samples were analyzed extensively with a total of 70 acutely infected patient serum samples. Eight of these clones failed to react with any of the patient sera. Each of these eight clones, when tested individually with 92 serum samples from the immune group, reacted wi...