TY  - JOUR
A1  - Beiss, Veronique
A1  - Spiegel, Holger
A1  - Boes, Alexander
A1  - Scheuermayer, Matthias
A1  - Reimann, Andreas
A1  - Schillberg, Stefan
A1  - Fischer, Rainer
T1  - Plant expression and characterization of the transmission-blocking vaccine candidate PfGAP50
JF  - BMC Biotechnology
N2  - Background: Despite the limited success after decades of intensive research and development efforts, vaccination still represents the most promising strategy to significantly reduce the disease burden in malaria endemic regions. Besides the ultimate goal of inducing sterile protection in vaccinated individuals, the prevention of transmission by so-called transmission blocking vaccines (TBVs) is being regarded as an important feature of an efficient malaria eradication strategy. Recently, Plasmodium falciparum GAP50 (PfGAP50), a 44.6 kDa transmembrane protein that forms an essential part of the invasion machinery (glideosome) multi-protein complex, has been proposed as novel potential transmission-blocking candidate. Plant-based expression systems combine the advantages of eukaryotic expression with a up-scaling potential and a good product safety profile suitable for vaccine production. In this study we investigated the feasibility to use the transient plant expression to produce PfGAP50 suitable for the induction of parasite specific inhibitory antibodies. 
Results: We performed the transient expression of recombinant PfGAP50 in Nicotiana benthamiana leaves using endoplasmatic reticulum (ER) and plastid targeting. After IMAC-purification the protein yield and integrity was investigated by SDS-PAGE and Western Blot. Rabbit immune IgG derived by the immunization with the plastidtargeted variant of PfGAP50 was analyzed by immune fluorescence assay (IFA) and zygote inhibition assay (ZIA). PfGAP50 could be produced in both subcellular compartments at different yields IMAC (Immobilized Metal Affinity Chromatography) purification from extract yielded up to 4.1 mu g/g recombinant protein per fresh leaf material for ER-retarded and 16.2 mu g/g recombinant protein per fresh leave material for plasmid targeted PfGAP50, respectively. IgG from rabbit sera generated by immunization with the recombinant protein specifically recognized different parasite stages in immunofluorescence assay. Furthermore up to 55 % inhibition in an in vitro zygote inhibition assay could be achieved using PfGAP50-specific rabbit immune IgG. 
Conclusions: The results of this study demonstrate that the plant-produced PfGAP50 is functional regarding the presentation of inhibitory epitopes and could be considered as component of a transmission-blocking malaria vaccine formulation.
KW  - PFS25
KW  - plastid targeting
KW  - plant-made vaccines
KW  - agroinfiltration
KW  - gametes
KW  - sexual stage
KW  - plasmodium falciparum
KW  - membrane
KW  - antibodies
KW  - immunization
KW  - RTS,S/AS01 malaria vaccine
KW  - recombinant proteins
KW  - cost-effectiveness
KW  - purification
Y1  - 2015
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-137327
VL  - 15
IS  - 108
ER  -