THIOREDOXIN FUSION HIV-1 PROTEASE COEXPRESSION SYSTEM FOR PRODUCTION OF SOLUBLE HUMAN IL6 IN ESCHERICHIA-COLI CYTOPLASM

In this paper, thioredoxin (TRX) fusion expression system has been mod ified to produce soluble human IL6 (hIL6) without TRX moiety in E. col i cytoplasm. A novel TRX gene fusion vector was developed that contain ed at the 3'-end of TRX gene a short DNA sequences encoding a linker p eptide '-GSGSGVSQNYPIVQHHHHHH-', serving not only as a specific HIV-1 protease site but also providing six contiguous histidine (His) residu es to foreign proteins. The cDNA for hIL6 was cloned into this vector resulting in plasmid pTRX@HISIL6. The cDNA for the HIV-1 protease has been cloned into another compatible plasmid pHMM2, resulting in plasmi d pHMM2-PR. Both plasmids were transformed into E.coli strain GI724, a nd when induced for expression of both proteins, the correct processin g of TRX@HISIL6 was obtained, producing hIL6 with Hiss-tag at the N te rminus named HISIL6. A fraction of HISIL6 was found in soluble form an d could be purified to homogeneity by Ni-NTA Superflow and ion-exchang e chromatography. The biological activity of purified HISIL6 was measu red by MTT method in an IL-6-dependent cell line 7TD1 to be 2.1 x 10(8 ) unit/mg.