CAJ | 학술논문

目的:构建 GFE-1多肽与重组人肿瘤坏死子α(rmhTNF-α)融合蛋白(GFE-1-rmhTNF),研究该融合蛋白的体外活性和体内分布.方法:利用基工程方法,将人工合成的编码 GFE-1的寡核苷酸片段连接在 rmhTNF-α序列的3'端,转入大肠杆菌中诱导表达,采用 Q-Sepharose FF 离子层析柱和 SP-Sepharose FF 阳离子层析柱纯化蛋白,SDS-PAGE 和 Western 印迹鉴定,测定该融合蛋白的体外活性,观察其在小鼠体内的分布情况.结果:构建了融合蛋白 GFE-1-rmhTNF,并在大肠杆菌中获得高效表达.体外活性实验显示,GFE-1-rmhTNF 对 L929细胞有明显的杀伤活性;体内分布实验证实,GFE-1-rmhTNF 在小鼠肺组织的富集高肝肾组织.结论:构建了融合蛋白 GFE-1-rmhTNF,可显著杀伤 L929细胞并特异性富集小鼠肺组织.
목적:구건 GFE-1다태여중조인종류배사자α(rmhTNF-α)융합단백(GFE-1-rmhTNF),연구해융합단백적체외활성화체내분포.방법:이용기공정방법,장인공합성적편마 GFE-1적과핵감산편단련접재 rmhTNF-α서렬적3'단,전입대장간균중유도표체,채용 Q-Sepharose FF 리자층석주화 SP-Sepharose FF 양리자층석주순화단백,SDS-PAGE 화 Western 인적감정,측정해융합단백적체외활성,관찰기재소서체내적분포정황.결과:구건료융합단백 GFE-1-rmhTNF,병재대장간균중획득고효표체.체외활성실험현시,GFE-1-rmhTNF 대 L929세포유명현적살상활성;체내분포실험증실,GFE-1-rmhTNF 재소서폐조직적부집고간신조직.결론:구건료융합단백 GFE-1-rmhTNF,가현저살상 L929세포병특이성부집소서폐조직.
Objective: To construct GFE-1-rmhTNF fusion protein and study its bioactivity in vitro and distribu?tion in vivo. Methods: The synthetic GFE-1 oligonucleotide fragment was linked to the 3' terminal of the se?quence encoding rmhTNF-α and expressed in E.coli DH5α. The fusion protein was purified by ion-exchange chro?matography and identified by SDS-PAGE and Western blot. Bioactivity in vitro and distribution in vivo of the fu?sion protein were further analysed. Results: GFE-1-rmhTNF was constructed and expressed in E.coli efficiently. In vitro, its significant cytotoxic activity in L929 cells was observed. In vivo, GFE-1-rmhTNF was dramatically en?riched in the lung tissue of mice than the liver and kidney tissue. Conclusion: We successfully constructed GFE-1-rmhTNF fusion protein. It could kill L929 cells efficiently and specifically enriched in the lung tissue of mice.