A novel gigaporous GSH affinity medium for high-speed affinity chromatography of GST-tagged proteins

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• 作者：Yongdong Huang ; Rongyue Zhang ; Juan Li ; Qiang Li ; Zhiguo Su ; Guanghui Ma
• 关键词：GSH affinity chromatography ; Gigaporous polystyrene microspheres ; High-speed chromatography ; GST-tagged proteins
• 刊名：Protein Expression and Purification
• 出版年：March, 2014
• 年：2014
• 卷：95
• 期：Complete
• 页码：84-91
• 全文大小：1804 K

文摘

Novel GSH-AP (phenoxyl agarose coated gigaporous polystyrene, Agap-co-PSt) microspheres were successfully prepared by introducing GSH ligand into hydrophilic AP microspheres pre-activated with 1,4-butanediol diglycidyl ether. The gigaporous structure and chromatographic properties of GSH-AP medium were evaluated and compared with commercial GSH Sepharose FF (GSH-FF) medium. The macropores (100-500 nm) of gigaporous PSt microspheres were well maintained after coating with agarose and functionalized with GSH ligand. Hydrodynamic experiments showed that GSH-AP column had less backpressure and plate height than those of GSH-FF column at high flow velocity, which was beneficial for its use in high-speed chromatography. The presence of flow-through pores in GSH-AP microspheres also accelerated the mass transfer rate of biomolecules induced by convective flow, leading to high protein resolution and high dynamic binding capacity (DBC) of glutathione S-transferase (GST) at high flow velocity. High purity of GST and GST-tagged recombinant human interleukin-1 receptor antagonist (rhIL-1RA) were obtained from crude extract with an acceptable recovery yield within 1.5 min at a velocity up to 1400 cm/h. GSH-AP medium is promising for high-speed affinity chromatography for the purification of GST and GST-tagged proteins.