title: The construction of a bifunctional fusion protein consisting of SEC2 and EGFP.
abstract: The aim of this study was to construct a bifunctional fusion protein consisting of staphylococcal enterotoxin C2 (SEC2) and enhanced green fluorescent protein (EGFP). We inserted EGFP and SEC2 fragments into the pET-28a(+) vector to create the expression plasmid vector, pET-28a(+)-SEC2-EGFP, using a two-step method. After verification of the plasmid, successful isolation of the fusion protein, SEC2-EGFP, was achieved by Ni+-affinity chromatography. Fluorescence microscopy, methylthiazol tetrazolium, and flow cytometry assays demonstrated that the constructed fusion protein not only retained the fluorescence signal of EGFP but also exhibited SEC2 bioactivity. Therefore, SEC2-EGFP is a promising tool for the study of the detailed temporal and spatial distributions of SEC2 in cells. Future studies with this vector may help uncover novel therapeutic strategies to treat or manage SEC2-associated diseases and be a new clinical tool for exploiting SEC2 in immunotherapy.
authors.affiliation: Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang, People's Republic of China; Department of Life Science and Technology, Xinxiang Medical University, Xinxiang, People's Republic of China.
authors.forename: Yanli
authors.initials: Y
authors.lastname: Liu
authors.forename: Mingkai
authors.initials: M
authors.lastname: Xu
authors.forename: Xu
authors.initials: X
authors.lastname: Li
authors.forename: Jian
authors.initials: J
authors.lastname: Sun
authors.forename: Chenggang
authors.initials: C
authors.lastname: Zhang
authors.forename: Huiwen
authors.initials: H
authors.lastname: Zhang
mesh_terms: D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004768:Enterotoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli; D005260:Female; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008214:Lymphocytes; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids; D015202:Protein Engineering / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D004768:Enterotoxins; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; C034959:enterotoxin C, staphylococcal
keywords: T cell activation; enhanced green fluorescent protein; fusion protein; staphylococcal enterotoxin C2; superantigen