Construction of bifunctional fusion proteins consisting of duck BAFF and EGFP.
We constructed fusion proteins consisting of fluorescence-enhanced green fluorescent protein (EGFP) and soluble domain of duck B-cell-activating factor of the TNF family (dsBAFF). The soluble EGFP/dsBAFF was efficiently expressed in Escherichia coli BL 21 (DE3) and was purified in milligram amounts using metal chellate affinity chromatography. The fusion protein exhibited similar fluorescence spectra with free EGFP and promoted the survival of duck bursal B cells in vitro as well as dsBAFF. EGFP/dsBAFF has shown specific binding to duck BAFF receptors positive-cells and the stained cells could be analyzed with flow cytometry. Thus, the fusion protein represents a readily obtainable source of biologically active dsBAFF that may prove useful in further studies on duck BAFF and its receptors.
Dan
Wen-Bing
WB
Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing 210097, China. danwenbing@126.com
Zhang
Chao
C
Guan
Zheng-Bing
ZB
Zhang
Shuang-Quan
SQ
D000818:Animals; D053264:B-Cell Activating Factor / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D004372:Ducks; D004926:Escherichia coli / Q000235:genetics; D005456:Fluorescent Dyes; D049452:Green Fluorescent Proteins / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics
D053264:B-Cell Activating Factor; D005456:Fluorescent Dyes; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins