Expression, purification of recombinant Luxi yellow cattle IFN-alpha fusion protein and its antiviral activities.
- Author:
Yong-Hong ZHANG
1
;
Chang-Fa WANG
;
Shao-Hua YANG
;
Yun-Dong GAO
;
Hong-Mei WANG
;
Jing-Peng LI
;
Ji-Feng ZHONG
Author Information
1. Research Center of Dairy Cattle Science, Shandong Academy of Agricultural Science, Jinan 250100, China.
- Publication Type:Journal Article
- MeSH:
Amino Acid Sequence;
Animals;
Antiviral Agents;
isolation & purification;
Base Sequence;
Cattle;
genetics;
Escherichia coli;
genetics;
metabolism;
Interferon-alpha;
biosynthesis;
genetics;
isolation & purification;
Molecular Sequence Data;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
isolation & purification;
Rotavirus;
drug effects;
Sequence Analysis
- From:
Chinese Journal of Biotechnology
2007;23(4):730-734
- CountryChina
- Language:Chinese
-
Abstract:
Interferon a gene was cloned from genomic DNA of Chinese Luxi yellow cattle by PCR, and the PCR product was inserted into vector pET32a( + ) to make a recombinant plasmid pET32a( + )/BoIFN-alpha. The expression of BoIFN-alpha in Escherichia coli was induced by addition of IPTG. Sequence analysis showed that the Chinese Luxi yellow cattle IFN-alpha gene is composed of 498 nucleotides, encoding a mature polypeptide of 166 amino acids. Compared with other BoIFN-alpha subtypes, it shares the highest identity of 97.6% to the C-subtype. SDS-PAGE results showed that recombinant proteins were expressed in inclusion bodies in Escherichia coli with molecular weight of 40 kD and the recombinant proteins accounted for 26.7% of the whole proteins.The expressed product was purified by affinity chromatography with immobilized nickel chelating NTA (Ni-NTA) and its antiviral activities were tested on MDBK/VSV cell system. Its antiviral activities were 5 x 10(5) u/mg on MDBK/VSV cell system. The results showed that the expression plasmid was successfully constructed and BoIFN-alpha C2 protein was expressed in Escherichia coli. Moreover the purification had good effects on antiviral activities.
