重组人r干扰素的纯化及其N端氨基酸序列分析

Purification and N-terminal Amino Acid Sequencing of Recombinant Human Interferon Gamma

本文对两个重组人γ干扰素高效表达株pIFN-γ及PBVIFN-γ的表达产物进行了纯化并对纯化的γ干扰素进行了活性鉴定及N端氨基酸序列分析。采用连续沉淀的方法对高表达菌株pBVIFN-γ及低表达菌株pIFN-γ,裂解液进行初步纯化,然后应用单克隆抗体亲和层析方法进行纯化,分别可纯化14倍与933倍,均达电泳纯,回收率分别为25％和30％,比活性达7.56×10~7U/mg蛋白。SDS-PAGE电泳上γ干扰素分子量约17.5kD。测定了纯化的γ干扰素N末端19个氨基酸序列,与由其DNA序列推导的氨基酸序列完全一致,确认了本研究所表达、纯化的γ干扰素达到了较高纯度。本方法为γ干扰素的批量生产奠定了基础。

Recombinant human interferon gamma (rHu IFN-γ) produced in E. co-li carrying plasmid pBVIFN-γ or pIFN-γ was purified to homogeneity with recoveries of 30% and 25% respectively, The purified rHu IFN-y appeared on SBS-PAGE electrophoresis as a single band of MW 17.5. kD The specific activity of the purified r huIFN-γ reached 7.56 × 107IU/mg protein. N terminal amino acid sequencing showed that the protein expressed and purified was rHuIFN-γ.