Long-acting Recombinant Fusion Protein Drugs Hsa - Gs - Stnfr1 And Stnfr1 - Hsa In Yeast Secretory Expression In The System Research

Tumor necrosis factorα(TNFα) functions through its two cell receptors, TNFR1 and TNFR2. The extracellular parts of TNFRs may fall off by proteolytic processing, and form soluble TNF receptors (sTNFR1 and sTNFR2), which play an important regulatory role in the biological effects of TNF-α. The natural sTNFR monomer has short half-life in vivo, and its affinity of combination with TNFa is very low.We introduced human serum albumin (HSA) fusion technology. Taking advantage of HSA’s stable structure, long half-life and large molecular weight, we fused the entire HSA strand to N and C terminus of sTNFR1 respectively, and obtained recombinant fusion proteins HSA-GS-sTNFRl and sTNFR1-HSA. The fusion protein HSA-GS-sTNFR1 contains a flexible short peptide (GGGGS) 2. We expressed the fusion proteins in yeast GS115, and obtained strains with high level expression of secreted HSA-GS-sTNFR1 and sTNFR1-HSA. We established a laboratory purification process. Through the cation exchange column and the gel chromatography, the purity of fusion proteins could be over 90%. The purified protein HSA-GS-sTNFR1 and sTNFR1-HSA could neutralize the cytotoxicity of TNFa in L929 cells, while the activity of sTNFR1-HSA is 3-4 times higher than that of HSA-GS-sTNFR1.The present research explored the expression, purification and biological activity of the recombinant fusion protein HSA-GS-sTNFR1 and sTNFR1-HSA, which laid the foundation for future development of long-acting TNFR protein drugs.