Tat短肽跨膜递送作用的研究(I)——模型蛋白Tat-GFP在毕赤酵母中的表达与纯化

Study on the transduction activity of Tat oligopeptide (I)——expression and purification of the model protein Tat-GFP in P.pastoris

为探讨Tat短肽的跨膜递送作用,利用DNA重组技术在毕赤酵母表达系统中表达融合蛋白Tat-GFP,通过硫酸铵沉淀、SephadexG-75凝胶色谱和POROS-20HQ离子交换色谱分离纯化该融合蛋白.表达和纯化了分子量约为29kD的融合蛋白Tat-GFP,在经融合蛋白作用的细胞内检测到融合蛋白的存在.这个结果可望为外源性蛋白进行细胞内治疗提供一种新的工具.

In order to investigate the transduction mechanism of Tat oligopeptide, the fusion protein Tat-GFP was expressed in P.pastoris expression system, and then purified into electrophoritic purity by precipitation of ammonium sulfate, Sephadex G-75 gel filtration chromatography and POROS-20HQ anion-exchange chromatography. Results showed that Tat-GFP fusion protein with molecular weight of 29kD was successly expressed and purified. After incubation with the liver cells, the purified Tat-GFP was found in most of cells. It is promising to provide a novel approach for the intracellular treatment with exogenous protein.