Synthetic gene construct expressing a repeated and highly immunogenic epitope of the Plasmodium falciparum antigen Pf155.

Åslund, Lena; Sjölander, Anders; Wahlgren, Mats; Wåhlin, B; Ruangjirachuporn, Wipaporn; Berzins, Klavs; Wigzell, Hans; Perlmann, Peter; Pettersson, Ulf

doi:10.1073/pnas.84.5.1399

Cited by 11 publications

(7 citation statements)

References 13 publications

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“…Injection into rabbits of carrier protein-conjugated peptides representing the 3' repeats of Pfl55/RESA or of fusion proteins encoded by corresponding gene constructs (Aslund et al 1987, St&hl et al 1989 gives rise to antibodies with properties very similar to those of the human antibodies discussed above. It is evident, therefore, that these sequences may also be a suitable basis for an immunogen in a subunit malaria vaccine.…”

Section: Discussionmentioning

confidence: 99%

“…These studies are primarily based on analyses of blood samples from adult donors living in an area of Northern Liberia where P. falciparum malaria is holoendemic and perennial (for details see Bjorkman et al 1985, Petersen et al 1989a). The results are laying the ground for the selection of B-cell epitopes for a Pfl 55/ RESA-based vaccine immunogen produced by recombinant gene technology (Aslund et al 1987, St4hl et al 1989. They are also an important basis for "epitope specific" sero-epidemiological studies (Bjorkman et al 1989, Petersen et al 1989a which are a prerequisite for future vaccine trials.…”

Section: Introductionmentioning

confidence: 99%

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Dissection of the human antibody response to the malaria antigen Pf155/RESA into epitope specific components

Perlmann

Berzins

et al. 1989

Immunological Reviews

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The development of vaccines is presently receiving major attention in malaria research. As it is not possible to base malaria vaccines on the use of killed or attenuated organisms, the vaccines which are being developed are subunit vaccines in which the immunogens consist of defined parasite antigens or antigenic fragments. Since protective immunity to malaria involves both antibody-dependent and antibody-independent mechanisms, the immunogens in a subunit vaccine must have the capacity to induce relevant B- and T-cell responses in the majority of vaccinees. In turn, this requires good knowledge of these responses in humans who have acquired immunity through natural infection. In this paper we have summarized our recent work on the dissection into epitope-specific components of the human antibody response to the Plasmodium falciparum antigen Pf155/RESA, a recognized candidate for a vaccine against the asexual blood stages of this parasite. Epitope mapping of the antigen by means of short synthetic peptides led to the identification in several molecular regions of short amino acid sequences constituting linear and probably immunodominant B-cell epitopes. The antigenically most active region was located in the C-terminus of the molecule. This region, which consists of approximately 40 related, 4- or 8-amino acid long repeats, induced higher antibody concentrations in a larger number of malaria-immune donors than any of the other regions. A large fraction of these antibodies bound to short synthetic peptides representing the major repeat motifs of Pf155/RESA. Although these repeats are made up of closely related amino acid sequences, the antibody response to them was highly polyclonal, indicating the presence of several linear and probably also conformational epitopes which gave rise to a variety of cross-reacting as well as monospecific antibodies. Further analysis revealed that the levels of antibodies differing in specificity and/or avidity for different peptides varied independently of each other in individual donors. In an area (Liberia) where malaria transmission is holoendemic and perennial, these antibody profiles remained constant when individual donors were followed over several years. Since the C-terminal repeat region of Pf155/RESA is conserved in different P. falciparum strains, the results reflect differences in the genetic regulation of epitope-specific host responses rather than antigenic differences between infecting parasites. In donors living in an area with high but seasonal malaria transmission, antibody levels usually drop to lower levels when there is no transmission.(ABSTRACT TRUNCATED AT 400 WORDS)

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Dissection of the human antibody response to the malaria antigen Pf155/RESA into epitope specific components

Perlmann

Berzins

et al. 1989

Immunological Reviews

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“…The construction, expression, and purification of the fusion proteins has been described in detail elsewhere (21). Briefly, the synthetic gene fragment encoding a tetramer of the C-terminal octapeptide repeat EENVEHDA of the P. falciparum antigen Pfl55/ RESA was cut out from a construction in the expression vector pATH 11 (1) and inserted into plasmid pEZZT308 or pBlB2mpl8 (15). The resulting expression vectors, pEZZM1 and pBlB2M1, encode the octapeptide repeats and either a divalent synthetic IgG-binding domain derived from staphylococcal protein A or the HSA-binding domains of streptococcal protein G, respectively.…”

Section: Methodsmentioning

confidence: 99%

“…Immunoblotting. Parasite extracts were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions, electrophoretically transferred to nitrocellulose, and probed with rabbit serum as previously described (1,4).…”

Section: Methodsmentioning

confidence: 99%

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Immunogenicity and antigenicity in rabbits of a repeated sequence of Plasmodium falciparum antigen Pf155/RESA fused to two immunoglobulin G-binding domains of staphylococcal protein A

et al. 1990

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A synthetic gene encoding a tetramer of the repeated subunit EENVEHDA of the Plasmodium falciparum antigen Pfl55/RESA was expressed in a dual-expression system. The resulting fusion proteins, designated ZZ-M1 and BB-M1, comprised the EENVEHDA repeats and either two immunoglobulin G-binding domains from staphylococcal protein A or the human serum albumin-binding domains from streptococcal protein G, respectively. The soluble fusion proteins were affinity purified to homogeneity in one-step procedures. ZZ-M1 was used for immunization of rabbits. The rabbit antisera reacted with BB-M1 in an enzyme-linked immunosorbent assay and with Pf155/RESA in immunofluorescence of infected erythrocytes and immunoblotting. Inhibition studies revealed that the antibodies mainly recognized epitopes formed by two or more EENVEHDA subunits and were remarkably specific for Pfl55/RESA. Importantly, the antibodies also inhibited P. falciparum merozoite reinvasion in vitro efficiently, indicating that they reacted with biologically important epitopes exposed on the native antigen. Immunization with Freund complete adjuvant resulted in high levels of specific immunoglobulin G antibodies over a 1-year period, whereas the antibody response obtained after immunization without adjuvant was generally weaker, immunoglobulin G and M mediated, and not sustained for longer periods. However, these titers were restored after booster injection. Taken together, the results support the usefulness of recombinant gene constructs of this type as immunogens for malaria vaccines.

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Molecular and cellular biology of malaria

Braun

1988

BioEssays

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SummaryThanks to the extensive use of recombinant DNA technology and immunological methods, much insight into cellular functions of the human malaria parasite Plasmodium falciparum has been gained since it was learnt ten years ago how to grow this organism in culture. The amino acid sequence of over a dozen surface proteins of the parasite and of several proteins the parasite excretes into its most important host cell, the erythrocyte, have been determined. Interestingly many of these proteins show blocks of repeated amino acids. Several proteins have been shown to be involved in specific aspects of the complex hostparasite interaction, such as penetration of host cells or increased stickiness of infected red blood cells in the blood vessels. Some of the proteins described here may be protective antigens and may become 'important in vaccine development.

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Synthetic gene construct expressing a repeated and highly immunogenic epitope of the Plasmodium falciparum antigen Pf155.

Cited by 11 publications

References 13 publications

Dissection of the human antibody response to the malaria antigen Pf155/RESA into epitope specific components

Dissection of the human antibody response to the malaria antigen Pf155/RESA into epitope specific components

Immunogenicity and antigenicity in rabbits of a repeated sequence of Plasmodium falciparum antigen Pf155/RESA fused to two immunoglobulin G-binding domains of staphylococcal protein A

Molecular and cellular biology of malaria

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