Preparation and bioactivity evaluation of streptavidin-tagged human interleukin-15 fusion protein.
- Author:
Hua SU
1
;
Yan-Li CHEN
;
Su-Yun CHEN
;
Bo WEN
;
Hong-Sheng YU
;
Zhi-Ming HU
;
Ji-Min GAO
Author Information
- Publication Type:Journal Article
- MeSH: Cancer Vaccines; genetics; immunology; Escherichia coli; genetics; metabolism; Genetic Vectors; Humans; Interleukin-15; biosynthesis; genetics; Lymphocyte Activation; drug effects; Recombinant Fusion Proteins; biosynthesis; genetics; immunology; Streptavidin; biosynthesis; genetics
- From: Journal of Southern Medical University 2009;29(3):397-401
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo obtain streptavidin-tagged human interleukin-15 (SA/hIL15) fusion protein and evaluate its bioactivity.
METHODSpET24a-6His-SA-hIL-15 and pET32a-hIL-15-SA-6His plasmids were constructed and expressed in BL 21(DE3) host bacteria to generate the fusion protein. The recombinant fusion protein IL-15/SA was purified using Ni-NTA affinity chromatography and refolded, and the efficiency of surface modification of the fusion protein on biotinylated cells was examined by fluorescence-activated cell sorting. CCK-8 method was used to evaluate the effect of IL-15/SA fusion protein in inducing the proliferation of human peripheral-blood lymphocyte (PBL) cells stimulated by PHA.
RESULTSThe recombinant SA-hIL-15 and hIL15-SA fusion proteins were highly expressed in BL21(DE3) at about 20% of the total bacterial proteins. The purified hIL15-SA fusion protein exhibited a bifunctionality by promoting the proliferation of PBL cells activated by PHA and high-affinity binding to biotinylated cell surface mediated by SA, with a cell surface modification efficiency exceeding 95%. SA-hIL-15 showed a 4-fold higher hIL15 bioactivity than hIL15-SA.
CONCLUSIONSA/hIL-15 bifunctional fusion protein has been successfully obtained to facilitate the future development of hIL-15-surface-modified cancer cell vaccine.
