Purification of VP3 protein of infectious bursal disease virus using nickel ion-immobilized regenerated cellulose-based membranes.
In this study, hexa-histidine tagged VP3 protein of infectious bursal disease virus (IBDV) was purified using immobilized metal ion affinity technique from the fermentation of Escherichia coli BL21 (DE3) containing a recombinant plasmid with a VP3 gene. The purification efficiencies of VP3 protein (TVP3 and DeltaTVP3) using Ni(2+)-NTA commercial agarose gels and Ni(2+)-IDA regenerated cellulose-based membranes at 4 degrees C were compared. A good washing condition for removing most impurity proteins was found as 20 mM NaH(2)PO(4), 500 mM NaCl, 40 mM imidazole, pH 7.8, whereas an efficient elution condition was 20 mM NaH(2)PO(4), 500 mM NaCl, 500 or 750 mM imidazole, pH 7.8. By applying these conditions to the flow experiments, similar recovery (86-88%) and purity (98-99%) for VP3 were obtained in both gel column (1 ml gel) and membrane cartridge (four membrane disks) under the flow rate of 1.7 ml/min for protein loading and 2.7 ml/min for protein elution. Regarding that the membrane process exhibited some advantages such as shorter residence time and lower cost, a better process efficiency in a large-scale system could be expected for the Ni(2+)-IDA membranes.
Hu
Hui-Ling
HL
Department of Chemical Engineering, National Chung Hsing University, Taichung 402, Taiwan.
Wang
Min-Ying
MY
Chung
Chiung-Hsuah
CH
Suen
Shing-Yi
SY
D002482:Cellulose / Q000737:chemistry*; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D008567:Membranes, Artificial; D009532:Nickel / Q000737:chemistry*; D015678:Viral Structural Proteins / Q000302:isolation & purification*
D008567:Membranes, Artificial; C478244:VP3 protein, infectious bursal disease virus; D015678:Viral Structural Proteins; D009532:Nickel; D002482:Cellulose