[Cloning and expression of human TNFR (P55)-IgG Fc fusion protein in eukaryotic cells].
OBJECTIVE
To express soluble biologically active TNF receptor protein in eukaryotic cells, which can inhibit cytotoxic activity of TNF.


METHODS
PCR amplified the extra-cellular region of TNF receptor P55 and IgG Fc gene. Then the two were linked through an oligomer encoding a thrombin-sensitive peptide linker and cloned into eukaryotic expression vector pcDNA3.1 (+). The eukaryotic expression plasmid pcDNA3. 1/TI was then transfected to the mammalian cell line COS7 and BHK. Using the G418 system, BHK cell clones were selected and can continuously secrete biological protein in large amount.


RESULTS
The expressed protein was fused with IgG Fc and secreted into the cell culture supernatant. It has good antigenicity and binding ability to TNF. It can also inhibit the cytotoxic activity of TNF on L929 cell.


CONCLUSIONS
The TNFR-IgG Fc fusion protein expressed in eukaryotic cells has biological activities of human TNF receptor P55.
An
N
N
State Key Laboratory of Molecular Virology and Genetic Engineering, Institute of Virology, Chinese Academy of Preventive Medicine, Beijing 100052, China.
Xu
C
C
Zhang
Z
Z
D000942:Antigens, Bacterial; D001426:Bacterial Proteins / Q000235:genetics*; D002478:Cells, Cultured; D005057:Eukaryotic Cells / Q000378:metabolism*; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000235:genetics*; D018124:Receptors, Tumor Necrosis Factor / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D007141:Immunoglobulin Fc Fragments; C097324:P55-L5 protein, Borrelia burgdorferi; D018124:Receptors, Tumor Necrosis Factor; D011993:Recombinant Fusion Proteins