人67kD层粘连蛋白受体在毕赤酵母中的表达及活性分析

Expression and Activity Analysis of Human 67kD Laminin Receptor in Pichia pastoris

摘要:

为实现人67kD层粘连蛋白受体（Human 67kD Laminin Receptor，67LR）蛋白的分泌表达，采用DNA重组技术将67LR cDNA片段插入分泌型酵母表达载体pPIC9K中，构建了相应的重组表达质粒pPIC9K-67LR并在GS115毕赤酵母菌株中表达，每升培养基经亲和层析可纯化目的蛋白12.56mg。纯化的目标蛋白能够与肺癌A549细胞竞争性结合其配体分子LN-1，具有相应的生物学活性，从而为深入研究人67LR的结构与功能奠定了基础。

Abstract:

To carry out the secretive expression of human 67kD laminin receptor (67LR)，recombinant expression plasmid pPIC9K-67LR was constructed by inserting of 67LR cDNA into yeast expression vector pPIC9K. The 67LR protein was expressed in Pichia pastoris after induced by methanol，and about 12.56mg electrophoresis purity 67LR could be obtained after the purification of 1L culture using affinity chromatograph column. In vitro competitive binding assay showed that target protein has an excellent biological activity. The successful expression of 67LR has placed a solid foundation for the research on structure and functions of 67LR.