Soluble Expression of Plasmodium falciparum Glutamate Dehydrogenase in Escherichia coli, and its Purification and Identification
- VernacularTitle:恶性疟原虫谷氨酸脱氢酶在大肠埃希菌中的可溶性表达、纯化和鉴定
- Author:
Yan LI
;
Yunshan NING
;
Wenqi DONG
;
Ming LI
- Publication Type:Journal Article
- Keywords:
Plasmodium falciparum;
glutamate dehydrogenase;
gene expression;
Western blotting
- From:
Chinese Journal of Parasitology and Parasitic Diseases
1987;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
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. Conclusion The soluble expression of GDH results in an integral three-dimensional structure epitope with high biological activity.
