关键词: 人源铁蛋白, 原核表达, 蛋白纯化, 可逆自组装

Abstract: In order to explore an efficient method for the preparation of reversible self-assembled active ferritin nanocage, we constructed prokaryotic expression system based on Escherichia coli for the recombinant human H-chain ferritin (BL21-pET3a-rHuHF) to express rHuHF efficiently in a fermentation tank. Three key parameters including stirring speed, air flux and induced expression time were optimized. A transmission electron microscope (TEM) and a light scattering instrument were used to characterize the nanostructure and particle size of rHuHF, respectively. The results showed that the optimum expression conditions were as follows: stirring speed 200 r/min, air flux 1.6 L/min, and expression time 9 h. According to the results of identification, ferritin existed in the soluble part of bacteria cells and had good self-assembly activity and monodisperse cage-like structure. The highest expression yield in 9 experiments was 290 mg/L, which was significantly higher than that in a shaking flask under the same conditions. The particle size of rHuHF was (11.23 ± 0.10) nm at pH 7 and (2.73 ± 0.10) nm at pH 2. This research can provide a scientific basis for the application of ferritinin nanometer-targeted delivery system.

Key words: ferritin, prokaryotic expression, protein purification, self-assembly