| Sichuan Da Xue Xue Bao Yi Xue Ban. 2016 Jan;47(1):14-8.
[Expression, Purification and Structure Prediction of Hum s1, a Major Allergen of
[Article in Chinese]
Liu Y, Sun XZ, Xu J, Wu YY.
OBJECTIVE: Hum s1, a major allergen of Humulus Scandens, was cloned, expressed
and purified. Its protein structure and function was predicted and analyzed,
which provided a foundation for further studies into the diagnostic and
therapeutic efficacy of the recombinant allergen.
METHODS: The target gene was amplified by PCR and sub-cloned into the expression
vector pET32a through Kpn I /Xho I site. The recombinant plasmid was transformed
into clone strain E. coli DH5α. The positive recombinant plasmid identified by
PCR was transformed into expression strain E. coli BL-21. The expressed fusion
protein was induced by IPTG, and purified using fast protein liquid
chromatography. The structure and function of the protein was predicted and
RESULTS: Prokaryotic expression plasmid pET32a-G2 was constructed successfully.
Hum s1 was expressed and purified. The purity of expressed fusion protein
exceeded 90%. It has three potential antigen epitopes and two EF-hand structural
domains. A three-dimensional model was successfully constructed. The recombinant
protein was proved to have immunological activity, with ELESA showing good
attachment to sera samples of patients who were allergic to Humulus Scandens.
CONCLUSION: Prokaryotic expression vector of Hum s1 was successfully constructed.
The recombinant protein was expressed and purified, with its epitope and
three-dimensional model being predicted by bioinformatics. The study provided a
basis for further development of recombinant vaccines.
PMID: 27062774 [Indexed for MEDLINE]