[Prokaryotic expression and characterization of two recombinant receptor-binding domain(RBD) proteins of human coronavirus NL63(HcoV-NL63)].
Bing Du Xue Bao
; 29(2): 106-11, 2013 Mar.
Article
en Zh
| MEDLINE
| ID: mdl-23757838
ABSTRACT
The receptor-binding domain(RBD) protein of HCoV-NL63 is a major target in the development of diagnostic assay and vaccine, it has a pivotal role in receptor attachment, viral entry and membrane fusion. In this study, we prepared 2 purified recombinant HCoV-NL63 RBD proteins using in E. coli system and identified the proteins by Western blotting. We first optimized codon and synthesized the RL (232-684aa)coding gene, then amplified the RL or RS(476-616aa) coding gene via PCR using different primers . The RL or RS coding gene was cloned into the pM48 expression vector fused with TrxA tag. The RBD (RL and RS) of HCoV-NL63 were expressed majorly as inclusion body when expressed in E. coli BL21pLys S under different conditions. The expressed products were purified by affinity chromatography then analyzed by SDS-PAGE and Western blotting. Our results showed that the recombinant RBD proteins were maximally expressed at 37 degrees C with 0. 8mM IPTG induction for 4h. RL or RS protein with 95 % purity was obtained and reacted positively with anti-sera from mice immunized with the recombinant vaccinia virus (Tiantan strain) in which HCoV-NL63 RL or RS protein was expressed. In conclusion, the purified recombinant RBD proteins(RL and RS)derived from E. coli were first prepared in China and they might provide a basis for further exploring biological role and vaccine development of HCoV-NL63.
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Base de datos:
MEDLINE
Asunto principal:
Proteínas del Envoltorio Viral
/
Infecciones por Coronavirus
/
Escherichia coli
/
Coronavirus Humano NL63
Tipo de estudio:
Prognostic_studies
Límite:
Animals
/
Humans
Idioma:
Zh
Revista:
Bing Du Xue Bao
Asunto de la revista:
VIROLOGIA
Año:
2013
Tipo del documento:
Article
País de afiliación:
China