Formation of high-capacity protein-adsorbing membranes through simple adsorption of poly(acrylic acid)-containing films at low pH.
Formation of high-capacity protein-adsorbing membranes through simple adsorption of poly(acrylic acid)-containing films at low pH.
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DOI：
10.1021/la300481e
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发表时间：
2012-05-01
期刊：
影响因子：
--
通讯作者：
Bruening ML
中科院分区：
文献类型：
--
作者：
Bhattacharjee S;Dong J;Ma Y;Hovde S;Geiger JH;Baker GL;Bruening ML
逐层吸附法是在多孔膜中引入离子交换位的一种简便方法。这项研究表明，吸附的聚（丙烯酸）（PAA），含膜在pH 3，而不是pH 5的蛋白质结合能力增加了3至6倍，这样的聚电解质改性膜。低吸附pH产生高密度的-COOH基团，其用作选择性结合标记蛋白质的金属离子络合物的共价固定的离子交换位点或点。当用次氮基三乙酸酯（NTA）-Ni 2+络合物官能化时，含有PAA/聚乙烯亚胺（PEI）/PAA膜的膜结合每cm 3膜93 mg组氨酸6-标记的（His-标记的）泛素。此外，这些膜从细胞提取物中分离His标记的COP 9信号体复合物亚基8，并显示>90%的His标记的泛素回收率。虽然用聚乙烯膜的改性通过简单地使聚乙烯溶液通过膜短至5分钟而发生，但在低pH沉积的情况下，这种膜的蛋白质结合能力与用聚合物刷改性的膜一样高，并且比市售IMAC树脂高2-3倍。此外，结合His标记的蛋白质的聚电解质改性膜的缓冲液渗透性是未改性膜的相应渗透性的约30%，因此蛋白质捕获可以在低压降下快速发生。即使在570 cm/h的溶液线速度下，用PAA/PEI/PAA改性的膜也表现出约40 mg/cm 3的溶菌酶动态结合容量（在10%穿透时的容量）。初步研究表明，这些膜在去热原条件（1 M NaOH）下是稳定的。
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- to 6-fold. The low adsorption pH generates a high density of –COOH 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)-Ni2+ complexes, membranes containing PAA/polyethyleneimine (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 2–3 fold higher than for commercially available 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).
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DOI：
10.1073/pnas.68.7.1393
发表时间：
1971-01-01
影响因子：
11.1
作者：
HARDMAN, KD;WOOD, MK;AINSWORTH, CF
通讯作者：
AINSWORTH, CF
DOI：
10.1038/nbt0788-779
发表时间：
1988-07-01
期刊：
BIO-TECHNOLOGY
影响因子：
--
作者：
BRANDT, S;GOFFE, RA;ZALE, SE
通讯作者：
ZALE, SE
影响因子：
3.9
作者：
Burke, SE;Barrett, CJ
通讯作者：
Barrett, CJ
影响因子：
2.9
作者：
BERGERS, JJ;VINGERHOEDS, MH;CROMMELIN, DJA
通讯作者：
CROMMELIN, DJA
影响因子：
2.8
作者：
Datta, Saurav;Bhattacharyya, D.;Toborek, M.
通讯作者：
Toborek, M.