[Preparation and bioactivity of anti-human red blood cell ScFv and CSFV E2 bifunctional fusion protein].
The aim of this study is to construct a bifunctional fusion protein, which can conjugate both human red blood cells and antibodies against classical swine fever virus (CSFV). We respectively amplified 2E8ScFv and mE2 genes from different recombinant vectors, in which 2E8ScFv gene is the single chain Fv gene against H antigen of human red blood cells, whereas mE2 gene is the main antigen coding region gene of CSFV E2 protein. We used overlap extension PCR to obtain an artificial fusion gene segment 2E8mE2 containing genes of Both 2E8ScFv and mE2, then ligated into the expression vector pET-DsbA and expressed in Escherichia coli BL21(DE3) PlysS host cells, after induced with IPTG the target fusion protein was successfully expressed and identified in inclusion bodies by SDS-PAGE and Western blotting. We purified the fusion protein and renatured it from inclusion bodies to obtain a native state of well biological activity. The Erythrocyte agglutination test results indicated that the fusion protein can conjugate both human red blood cells and antibodies of CSFV.
Qin
Shaomin
S
Guangxi Veterinary Research Institute, Nanning 530001, China.
Bai
Anbin
A
Wu
Jianmin
J
Liao
Wenjun
W
Yuan
Shuzhi
S
Hua
Jun
J
Guan
Zhongyi
Z
D004912:Erythrocytes / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D057127:Single-Chain Antibodies / Q000096:biosynthesis* / Q000235:genetics; D014759:Viral Envelope Proteins / Q000096:biosynthesis* / Q000235:genetics
D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D014759:Viral Envelope Proteins; C087086:glycoprotein E2, classical swine fever virus; C055966:immunoglobulin Fv