High-level expression of the malaria blood-stage vaccine candidate Plasmodium falciparum apical membrane antigen 1 and induction of antibodies that inhibit erythrocyte invasion.
Apical membrane antigen 1 (AMA-1) is a highly promising malaria blood-stage vaccine candidate that has induced protection in rodent and nonhuman primate models of malaria. Authentic conformation of the protein appears to be essential for the induction of parasite-inhibitory antibody responses. Here we have developed a synthetic gene with adapted codon usage to allow expression of Plasmodium falciparum FVO strain AMA-1 (PfAMA-1) in Pichia pastoris. In addition, potential N-glycosylation sites were changed, exploiting the lack of conservation of these sites in Plasmodium, to obtain high-level secretion of a homogeneous product, suitable for scale-up according to current good manufacturing procedures. Purified PfAMA-1 displayed authentic antigenic properties, indicating that the amino acid changes had no deleterious effect on the conformation of the protein. High-titer antibodies, raised in rabbits, reacted strongly with homologous and heterologous P. falciparum by immunofluorescence. In addition, purified immunoglobulin G from immunized animals strongly inhibited invasion of red blood cells by homologous and, to a somewhat lesser extent, heterologous P. falciparum.
Department of Parasitology, Biomedical Primate Research Centre, 2280 GH Rijswijk, The Netherlands.
Clemens H M
CH
Kocken
Chrislaine
C
Withers-Martinez
Martin A
MA
Dubbeld
Annemarie
A
van der Wel
Fiona
F
Hackett
Augusto
A
Valderrama
Michael J
MJ
Blackman
Alan W
AW
Thomas
D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology; D000953:Antigens, Protozoan / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D016054:DNA, Protozoan; D004912:Erythrocytes / Q000276:immunology / Q000469:parasitology; D015870:Gene Expression; D017780:Malaria Vaccines / Q000235:genetics / Q000276:immunology*; D016778:Malaria, Falciparum / Q000517:prevention & control*; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000235:genetics / Q000276:immunology*; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology*; D011817:Rabbits; D014614:Vaccines, Synthetic / Q000235:genetics / Q000276:immunology*
D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D016054:DNA, Protozoan; D017780:Malaria Vaccines; D008565:Membrane Proteins; D015800:Protozoan Proteins; D014614:Vaccines, Synthetic; C090077:apical membrane antigen I, Plasmodium