Formation of High-Capacity Protein-Adsorbing Membranes through Simple Adsorption of Poly(acrylic acid)-Containing Films at Low pH

详细信息    查看全文

• 作者：Somnath Bhattacharjee ; Jinlan Dong ; Yiding Ma ; Stacy Hovde ; James H. Geiger ; Gregory L. Baker ; Merlin L. Bruening
• 刊名：Langmuir
• 出版年：2012
• 出版时间：May 1, 2012
• 年：2012
• 卷：28
• 期：17
• 页码：6885-6892
• 全文大小：334K
• 年卷期：v.28,no.17(May 1, 2012)
• ISSN：1520-5827

文摘

Layer-by-layer polyelectrolyte adsorption is a simple, convenient method for introducing ion-exchange sites in porous membranes. This study demonstrates that adsorption of poly(acrylic acid) (PAA)-containing films at pH 3 rather than pH 5 increases the protein-binding capacity of such polyelectrolyte-modified membranes 3鈥?-fold. The low adsorption pH generates a high density of 鈭扖OOH groups that function as either ion-exchange sites or points for covalent immobilization of metal-ion complexes that selectively bind tagged proteins. When functionalized with nitrilotriacetate (NTA)鈥揘i²⁺ complexes, membranes containing PAA/polyethylenimine (PEI)/PAA films bind 93 mg of histidine₆-tagged (His-tagged) ubiquitin per cm³ of membrane. Additionally these membranes isolate His-tagged COP9 signalosome complex subunit 8 from cell extracts and show >90% recovery of His-tagged ubiquitin. Although modification with polyelectrolyte films occurs by simply passing polyelectrolyte solutions through the membrane for as little as 5 min, with low-pH deposition the protein binding capacities of such membranes are as high as for membranes modified with polymer brushes and 2鈥?-fold higher than for commercially available immobilized metal affinity chromatography (IMAC) resins. Moreover, the buffer permeabilities of polyelectrolyte-modified membranes that bind His-tagged protein are 30% of the corresponding permeabilities of unmodified membranes, so protein capture can occur rapidly with low-pressure drops. Even at a solution linear velocity of 570 cm/h, membranes modified with PAA/PEI/PAA exhibit a lysozyme dynamic binding capacity (capacity at 10% breakthrough) of 40 mg/cm³. Preliminary studies suggest that these membranes are stable under depyrogenation conditions (1 M NaOH).