Assembly and expression of a synthetic gene encoding the antigen Pfs48/45 of the human malaria parasite Plasmodium falciparum in yeast.
Pfs48/45 is an important transmission-blocking vaccine candidate antigen of the human malaria parasite Plasmodium falciparum. This study was aimed at synthesis of recombinant Pfs48/45 containing conformation-constrained epitopes of the native antigen in yeast. Since in the yeast Saccharomyces cerevisiae induction of gene-expression led to prematurely terminated transcripts, an entirely synthetic gene of higher GC content was assembled. Replacement of the AT rich natural gene by the synthetic gene relieved the observed premature transcription termination. Nevertheless, recombinant protein expression could not be detected. In contrast, in the yeast Pichia pastoris low levels of recombinant Pfs48/45 were produced upon induction of synthetic gene expression. The recombinant protein was shown to be disulphide-bridge constrained, but was not recognised by transmission-blocking antibodies and did not induce transmission-blocking sera in mice.
Milek
R L
RL
Department of Molecular Biology and Cell Biology, University of Nijmegen, Toernooiveld 1, 6525 ED Nijmegen, The Netherlands.
Stunnenberg
H G
HG
Konings
R N
RN
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016054:DNA, Protozoan / Q000235:genetics; D005813:Genes, Synthetic; D006801:Humans; D008288:Malaria / Q000469:parasitology; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics / Q000378:metabolism; D010963:Plasmodium falciparum / Q000235:genetics*; D015800:Protozoan Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D016054:DNA, Protozoan; D008562:Membrane Glycoproteins; D015800:Protozoan Proteins; C084847:pfs48-45 protein, Plasmodium falciparum