Preparation and bioactivity assay of mIL-4-SA fusion protein
- VernacularTitle:mIL-4-SA双功能融合蛋白的制备和活性测定
- Author:
Zhen ZHANG
;
Pingping FA
;
Jinlong LI
;
Zhiming HU
;
Jimin GAO
- Publication Type:Journal Article
- Keywords:
mouse intedeukin-4;
streptavidin;
fusion protein
- From:
Chinese Journal of Biochemical Pharmaceutics
2010;31(2):90-93
- CountryChina
- Language:Chinese
-
Abstract:
Purpose To prepare streptavidin-tagged mouse interleukin-4(mIL-4-SA)bifunctional fusion protein and to study on its bioactivity.Methods The mIL-4 gene was cloned by RT-PCR and cloned into pET21 vector to get mIL-4-SA-pET21 expression plasmid.The mIL-4-SA fusion protein was expressed in BL21 (DE3)host bacteria and purified through the Ni-NTA affinity chromatography and refolded by dilution and dialysis.The effect of mIL-4-SA fusion protein on mouse thymocytes proliferation was evaluated by MTY.Flow cytometric analysis was performed to detect the mIL-4-SA fusion protein on the biotinylated B16F10 tumor cells.Results The mIL-4-SA-pET21 vector was successful by constructed and the mIL-4-SA fusion protein was expressed in BL21(DE3)at about 35%of total bacterial proteins.The purity of mIL-4-SA Was about 95% through Ni-NTA.The mIL-4-SA fusion protein exhibited bifunctional activities,i.e.,stimulative effect for mouse thymocyte proliferation and SA-mediated high-affinity binding to biotinylated cell surfaces(anchoring modified rate Was about 96.69%).Conclusion The mIL-4-SA fusion protein was expected to be developed for the treatment of tumors.
