Fusion of two malaria vaccine candidate antigens enhances product yield, immunogenicity, and antibody-mediated inhibition of parasite growth in vitro.
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.
Pan
Weiqing
W
Department of Etiologic Biology, Second Military Medical University, Shanghai, China. malaria@guomai.sh.cn
Huang
Daqing
D
Zhang
Qingfeng
Q
Qu
Li
L
Zhang
Dongmei
D
Zhang
Xiaoli
X
Xue
Xiangyang
X
Qian
Feng
F
D000276:Adjuvants, Immunologic / Q000008:administration & dosage / Q000138:chemical synthesis / Q000235:genetics; D000818:Animals; D000913:Antibodies, Protozoan / Q000502:physiology*; D000953:Antigens, Protozoan / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology; D000981:Antiprotozoal Agents / Q000008:administration & dosage / Q000138:chemical synthesis / Q000276:immunology*; D002135:Calcium-Binding Proteins / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology; D006131:Growth Inhibitors / Q000008:administration & dosage / Q000138:chemical synthesis / Q000235:genetics / Q000276:immunology; D008253:Macaca mulatta; D017780:Malaria Vaccines / Q000008:administration & dosage / Q000138:chemical synthesis / Q000235:genetics* / Q000276:immunology; D008565:Membrane Proteins / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000254:growth & development* / Q000276:immunology; D011494:Protein Kinases / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology; D015800:Protozoan Proteins / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000138:chemical synthesis / Q000235:genetics* / Q000276:immunology; D013381:Subtilisins / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000138:chemical synthesis / Q000276:immunology*
D000276:Adjuvants, Immunologic; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D000981:Antiprotozoal Agents; D002135:Calcium-Binding Proteins; D006131:Growth Inhibitors; D017780:Malaria Vaccines; D008565:Membrane Proteins; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; C090077:apical membrane antigen I, Plasmodium; D011494:Protein Kinases; C079866:PfCPK protein, Plasmodium falciparum; D013381:Subtilisins; C119550:sub-2 protein, Plasmodium falciparum