[Soluble expression of Plasmodium falciparum glutamate dehydrogenase in Escherichia coli, and its purification and identification].
OBJECTIVE
To make soluble expression of Plasmodium falciparum (FCC1/HN) glutamate dehydrogenase (GDH) in Escherichia coli, purification and immunocompetence identification of the recombinant non-fusion GDH.


METHODS
The GDH gene was cloned into prokaryotic expression vector pET23 (a) to form recombinant expression vector pET23 (a)/GDH. pET23(a)/GDH was transformed into E. coli BL21 (DE3). Induced by IPTG (isopropyl-beta D-thiogalactoside), GDH was highly expressed in the supernatant after sonication. The soluble recombinant GDH was purified by Source-Q and Source-S chromatography. Enzyme-linked immunosorbent assay and Western blotting were carried out to identify the immunocompetence of the purified product.


RESULTS
SDS-PAGE analysis showed that the soluble GDH protein accounted for approximately 15% of the total bacterial protein. By two-step ion-exchange chromatography, the purity of GDH reached more than 90% and the GDH possessed high antigenicity.


CONCLUSIONS
The soluble expression of GDH results in an integral three-dimensional structure epitope with high biological activity.
Institute of Tropical Medicine, First Military Medical University, Guangzhou 510515, China.
Yan
Y
Li
Yun-shan
YS
Ning
Wen-qi
WQ
Dong
Ming
M
Li
D000818:Animals; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005969:Glutamate Dehydrogenase / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification; D007121:Immunocompetence; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification
D011994:Recombinant Proteins; D005969:Glutamate Dehydrogenase