基因工程法制备^(15)N标记的蛋白质样品

Preparation of 15 N labeled protein sample by gene engineering techonology

为了用多维核磁共振方法解析蛋白质溶液构象奠定基础，在用基因工程法使天花粉蛋白在大肠杆菌中获得高效表达的基础上，用１５Ｎ标记了这种蛋白质并通过亲和层析方法进行纯化得到电泳纯的蛋白样品；测定了此蛋白对中期妊娠小鼠的止孕效应；在核磁共振波谱仪上测得了此蛋白的１Ｈ－１５Ｎ异核相关谱（ＨＳＱＣ）。结果表明，上述方法可以用于制备１５Ｎ标记的蛋白质样品。

Using the advanced multidimension heteronuclear pulses and isotope labeled protein technique, nuclear magnetic resonance spectroscopy has become an important tool in analysis of the solution conformation of protein. In this report, on the basis of the high level expression of a protein trichosanthin in recombinant E.coli using DNA, 15 N was used to label the protein, the 15 N labeled trichosanthin was obtained by affinity chromatography on Ni NTA agarose. Terminating pregnant effect in mice showed that this recombinant protein had the same activity as natural trichosanthin. A 1 H 15 N heteronuclear single quantum coherence (HSQC) spectrum was obtained from an AM 500 NMR spectrometer, demonstrating that this method is suitable in preparing labeled protein sample for NMR.