Preparation of fusion protein between ZZ affinity peptide and alkaline phosphatase and its primary application
10.3969/j.issn.1000-484X.2015.09.014
- VernacularTitle:ZZ亲和肽-碱性磷酸酶融合蛋白的制备与初步应用
- Author:
Rumeng BAO
;
Shuangjia ZHAO
;
Min XUE
;
Hongming YANG
;
Jinbao TANG
- Publication Type:Journal Article
- Keywords:
ZZ affinity peptide;
Alkaline phosphatase;
Fusion protein;
Application
- From:
Chinese Journal of Immunology
2015;(9):1214-1216,1220
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct and express the fusion protein between ZZ affinity peptide and alkaline phosphatase and examine its biological activities.Methods:The alkaline phosphatase gene was cloned into pEZZ 18 vector containing ZZ peptide gene resulted in the pEZZ-AP recombinant vector.Then the vector was transformed and expressed in E.coli DH5α.And HisTrap affinity chromatography was employed to separate and purify the target protein.After analyzed by Western blot , ZZ-AP fusion protein was applied to immunocytochemistry as an alterative second antibody.Results:The result of SDS-PAGE showed that the fusion protein with a relative molecular weight was consistent with the theoretical values (62 kD).The fusion protein has rabbit IgG-binding ability and en-zymatic activity of alkaline phosphatase ,those were validated in Western blot;and it produced a good signal that was comparable in its staining pattern to that generated with goat anti-rabbit IgG-AP in immunocytochemistry.Conclusion: The ZZ-AP fusion protein was constructed successfully ,it has rabbit IgG-binding ability and enzymatic activity of alkaline phosphatase.We anticipate that the ZZ-AP fusion protein has a potential application in immunoassay field.
