猪白细胞介素-6在巴斯德毕氏酵母(Pichia pastoris)中的分泌表达

白细胞介素6 (Interleukin_6 ,IL_6 )是一种具有多种生物学效应的细胞因子,在疾病诊断与疫苗佐剂领域有广阔的应用前景。在本试验中,猪白细胞介素_6 (pIL_6 )的cDNA序列被克隆入甲醇酵母(Pichiapastoris)分泌表达载体pPIC9K中,并转化入P .pastorisGS115菌株。其重组菌株GS115 pPIC9K_IL6经1%甲醇诱导后,能分泌表达分子量约为2 4 5KD的重组蛋白,Westernblot确证为pIL_6。该酵母表达产物无N端糖基化修饰。用依赖IL6生长的B9细胞株检测提纯后的pIL_6 ,其生物学活性可达8×10 4 IU mg。

Expression of porcine interleukin-6 in Pichia pastoris

As a kind of pleiotropic cytokine, Interleukin-6 (IL-6) becomes more and more popular in the fields of disease diagnosis and candidate adjuvants. In this study, the porcine Interleukin-6 (pIL-6) cDNAs un-coding signal peptides was cloned into pPIC9K (the expression vector of Pichia pastoris) to construct the pPIC9K-IL6, which was further transformed into P. pastoris GS115 strain. After being induced by 1% methanol, the recombinant strain excreted a protein in supernate of culture, with molecular weight of 24.5KD approximately, which was proved to be pIL6 through SDS-PAGE and western blotting. This P. pastoris product wasn't modified by N-hyperglycosylation. After purified, pIL-6 could reach to 8 x 10(4) IU/mg of bioactivity unit by the cell proliferation of B9 assay. Through researching the characters of the recombinant protein pIL6, we have made a great progress on the study of rpIL6 as veterinary vaccine's adjuvant.