人cubilin蛋白功能片段原核表达系统的建立

Establishment of prokaryotic expression system of CUB7-8 fragment of hcubilin

目的构建编码白蛋白吞饮受体人cubilin(hcubilin)蛋白配体结合域CUB78基因片段的原核表达载体,诱导表达并鉴定表达产物。方法重组DNA技术构建原核表达载体pGEX hCUB78,IPTG诱导表达,SDS PAGE与免疫印迹法鉴定表达产物,Pulldown实验验证融合蛋白是否与白蛋白结合。结果双酶切和测序鉴定表明CUB78片段插入正确,诱导表达后获得分子量为52×103的融合蛋白,该融合蛋白可以被抗GST抗体识别,并且可以和白蛋白结合。结论CUB78基因片段编码的蛋白质能在原核细胞中正确表达,并且可以和白蛋白结合,这为进一步研究该功能域的结构与功能奠定了基础。

Objective To construct CUBT-8 fragment prokaryotic expression vector of humancubilin, the endocytic receptor for albumin and to induce expression and identify the expression product. Methods The expression vector pGEX-hCUB7-8 was constructed by recombinant DNA technology and the expressed protein was identified by SDS-PAGE and Western blotting after induction by IPTG, then the interaction between the fuprotein. Conclusion CUB7-8 protein can be expressed in E. coli. expression system and will combine with albumin. It is applicable for further investigation of cubilin.