Abstract
A Plasmodium falciparum chimeric protein 2.9 (PfCP-2.9) was constructed consisting of the C-terminal regions of two leading malaria vaccine candidates, domain III of apical membrane ag-1 (AMA-1) and 19-kDa C-terminal fragment of the merozoite surface protein 1 (MSP1). The PfCP-2.9 was produced by Pichia pastoris in secreted form with a yield of 2600 mg/L and approximately 1 g/L of final product was obtained from a three-step purification process. Analysis of conformational properties of the chimeric protein showed that all six conformational mAbs interacted with the recombinant protein were reduction-sensitive, indicating that fusion of the two cysteine-rich proteins retains critical conformational epitopes. PfCP-2.9 was found to be highly immunogenic in rabbits as well as in rhesus monkeys (Macaca mulatta). The chimeric protein induced both anti-MSP1-19 and anti-AMA-1(III) Abs at levels 11- and 18-fold higher, respectively, than individual components did. Anti-PfCP-2.9 sera from both rabbits and rhesus monkeys almost completely inhibited in vitro growth of the P. falciparum FCC1/HN and 3D7 lines when tested at a 6.7-fold dilution. It was shown that the inhibition is dependent on the presence of Abs to the chimeric protein and their disulfide bond-dependent conformations. Moreover, the activity was mediated by a combination of growth-inhibitory Abs generated by the individual MSP1-19 and AMA-1(III) of PfCP-2.9. The combination of the extremely high yield of the protein and enhancement of its immune response provides a basis to develop an effective and affordable malaria vaccine.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adjuvants, Immunologic / administration & dosage
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Adjuvants, Immunologic / chemical synthesis
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Adjuvants, Immunologic / genetics
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Animals
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Antibodies, Protozoan / physiology*
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Antigens, Protozoan / administration & dosage
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Antigens, Protozoan / genetics*
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Antigens, Protozoan / immunology
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Antiprotozoal Agents / administration & dosage
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Antiprotozoal Agents / chemical synthesis
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Antiprotozoal Agents / immunology*
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Calcium-Binding Proteins / administration & dosage
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Calcium-Binding Proteins / genetics*
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Calcium-Binding Proteins / immunology
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Growth Inhibitors / administration & dosage
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Growth Inhibitors / chemical synthesis
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Growth Inhibitors / genetics
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Growth Inhibitors / immunology
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Macaca mulatta
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Malaria Vaccines / administration & dosage
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Malaria Vaccines / chemical synthesis
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Malaria Vaccines / genetics*
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Malaria Vaccines / immunology
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Membrane Proteins / administration & dosage
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Membrane Proteins / genetics*
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Membrane Proteins / immunology
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Molecular Sequence Data
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Plasmodium falciparum / growth & development*
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Plasmodium falciparum / immunology
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Protein Kinases / administration & dosage
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Protein Kinases / genetics*
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Protein Kinases / immunology
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Protozoan Proteins / administration & dosage
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Protozoan Proteins / genetics*
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Protozoan Proteins / immunology
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Rabbits
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Recombinant Fusion Proteins / administration & dosage
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Recombinant Fusion Proteins / chemical synthesis
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Recombinant Fusion Proteins / genetics*
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Recombinant Fusion Proteins / immunology
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Subtilisins / administration & dosage
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Subtilisins / genetics
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Subtilisins / immunology
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Vaccines, DNA / administration & dosage
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Vaccines, DNA / chemical synthesis
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Vaccines, DNA / immunology*
Substances
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Adjuvants, Immunologic
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Antibodies, Protozoan
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Antigens, Protozoan
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Antiprotozoal Agents
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Calcium-Binding Proteins
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Growth Inhibitors
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Malaria Vaccines
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Membrane Proteins
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Protozoan Proteins
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Recombinant Fusion Proteins
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Vaccines, DNA
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apical membrane antigen I, Plasmodium
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Protein Kinases
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PfCPK protein, Plasmodium falciparum
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Subtilisins
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sub-2 protein, Plasmodium falciparum