人胰岛素原类似物(BKRA)基因三联体的构建与表达

Construction and Expression of Human Proinsulin Analog(BKRA)Gene Triplet in Eschreichia coli

将人胰岛素原类似物（ＢＫＲＡ）基因通过人工接头串联，形成基因三联体，接头部分的氨基酸序列为ＲＲＮＳＭ。该串联基因与表达载体ｐＥＴ２８ａ（＋）重组后，实现了在大肠杆菌中的高效融合表达，表达产物以包涵体形式存在，约占细菌总蛋白的３１％。以ＨｉＴｒａｐ凝胶进行亲和层析初步纯化，纯度可达９３％。放射免疫测定表明，纯化的融合蛋白具有胰岛素抗原活性，为进一步生产基因工程胰岛素打下了基础。

The human proinsulin analog(BKRA)gene molecules were linked tandemly with synthetic nucleotide linker coding RRNSM to form a triplet molecule.The BKRA gene triplet was cloned into pET 28a(+),an expression plasmid,and expressed in E.coli in a fusion protein fashion.The exressed proteins aggregated into inclusion body,and weigh about 31% of the total proteins of the total proteins of the expression bacteria.The expressed proteins are easily purified by HiTrap affinity matrix up to putity of 93%.Human insulin antigenicity of the purified fusion protein was detected by radioimmunoassay.