Purification and refolding of Escherichia coli-expressed recombinant human interleukin-2.
The expression of rhIL-2 (recombinant human interleukin-2) in bacteria results in the formation of insoluble inclusion-body aggregates. These aggregates were first solubilized under denaturing conditions (sodium phosphate buffer solution containing 8 M urea and 10 mM 2-mercaptoethanol) and then purified using IMAC (immobilized metal-ion-affinity chromatography). IMAC was used to capture rhIL-2. The protein was gradually refolded on the column by a gradient elution (8 M to 0 M urea) in the presence of 10% (v/v) glycerol. Glycerol was used to prevent protein aggregation during the refolding step. Using this method, rhIL-2 was collected at 97% purity and its activity was measured by the lymphocyte transformation test. The measured activity was identical with commercial human interleukin-2.
Esfandiar
Samaneh
S
Biotechnology Group, Chemical Engineering Department, Tarbiat Modares University, P.O. Box 14115-143, Tehran, Iran.
Hashemi-Najafabadi
Sameereh
S
Shojaosadati
Seyed Abbas
SA
Sarrafzadeh
Shokuh Aazam
SA
Pourpak
Zahra
Z
D002846:Chromatography, Affinity / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D002479:Inclusion Bodies / Q000378:metabolism*; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D009571:Nitrilotriacetic Acid / Q000031:analogs & derivatives / Q000737:chemistry; D009942:Organometallic Compounds / Q000737:chemistry; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012995:Solubility
D007376:Interleukin-2; D009942:Organometallic Compounds; D011994:Recombinant Proteins; C088321:nickel nitrilotriacetic acid; D009571:Nitrilotriacetic Acid