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采用金属螯合亲和层析技术纯化了N-末端组氨酸(His)标记的重组蛋白Ax Ce SD。以2000 ku的水溶性葡聚糖Dextran T2000为基质,亚氨基二乙酸(IDA)作为螯合剂,Cu2+做亲和配基制备了能特异性吸附重组蛋白His标记的水溶性葡聚糖亲和-超滤载体,并探讨了p H、离子浓度、吸附时间以及温度等因素对水溶性葡聚糖亲和-超滤载体吸附重组蛋白Ax Ce SD的影响。结果表明,在一定范围内,随着p H和温度的升高亲和-超滤载体对Ax Ce SD吸附量增加,而随着离子浓度的增加其对Ax Ce SD吸附量减少;亲和-超滤载体对Ax Ce SD的吸附在30 min内能够达到平衡。应用Langmuir方程拟合了等温条件下亲和-超滤载体对Ax Ce SD的吸附曲线,得到最大理论吸附量为125.15 mg/g,解离常数为3.259×10-6 mol/L,说明亲和-超滤载体对His标记的重组Ax Ce SD的吸附为以螯合作用为主的特异性吸附。
The N-terminal histidine (His) -labeled recombinant protein Ax Ce SD was purified by metal chelation affinity chromatography. Using 2000 ku water-soluble dextran Dextran T2000 as matrix and iminodiacetic acid (IDA) as chelating agent, Cu2 + as affinity ligand was prepared to specifically adsorb the recombinant protein His-tagged water-soluble dextran affinity- The effects of p H, ion concentration, adsorption time and temperature on the adsorption of recombinant protein Ax Ce SD by water-soluble dextran affinity-ultrafiltration were discussed. The results showed that the amount of adsorbed Ax Ce SD increased with the increasing of pH and temperature, while the amount of adsorbed Ax Ce SD decreased with the increase of ion concentration. The affinity- The adsorption of Ax Ce SD by the ultrafiltration carrier can be balanced within 30 min. The Langmuir equation was used to fit the adsorption curves of Ax-Ce SD to the affinity-ultrafiltration carrier under isothermal conditions. The maximum theoretical adsorption capacity was 125.15 mg / g and the dissociation constant was 3.259 × 10-6 mol / L, indicating that the affinity - Adsorption of the His-tagged recombinant Ax Ce SD by ultrafiltration vector is a specific chelate-based adsorption.