人釉原蛋白成熟肽基因的原核表达和纯化

Expression and purification of human amelogenin mature peptide in Escherichia coli

目的 建立人釉原蛋白（amelogenin，AMG）成熟肽基因原核表达和纯化的技术路线，获得纯化的人AMG成熟肽蛋白，以期为牙周炎的基础治疗提供依据。方法通过已构建并经鉴定的重组质粒pGEX-4T-1-AMG，转化B[21大肠杆菌，原核表达后利用谷胱甘肽-S-转移酶融合蛋白纯化系统（glutathioneS-transfe Fasefusion protein purificationsystem，GSTrapFF）亲和层析柱进行重组人AMG的纯化。结果十二烷基硫酸钠聚丙烯酰胺凝胶电泳图和蛋白质印迹法分析结果显示，获得纯化的相对分子质量为45000大小GST—AMG融合蛋白和19000大小的目的蛋白AMG。结论利用pGEX-4T-1-AMG-BL21体系成功获得了人AMG成熟肽基因在大肠杆菌中的原核表达和纯化。

Objective To establish the expression and purification route for human amelogenin mature peptide in Escherichia coli and abtain the purified amelogenin （AMG） mature peptide. Methods Recombined plasmid pGEX-4T-1-AMG was transformed to Escherichia coli BL21. After expression, AMG was purified with glutathione S-transferase fusion protein purification system （GSTrapFF） column. Results Sodium dodecyl sulfate polyacrylamide gel electrophoresis （SDS-PAGE） and Western blotting hybridization results showed that 45000 GST-AMG fusing protein and 19000 target AMG mature peptide were obtatined successfully. Condusions pGEX-4T-1-AMG-BL21 system is used successfully to express and purify human AMG mature peptide.