title: Formation of high-capacity protein-adsorbing membranes through simple adsorption of poly(acrylic acid)-containing films at low pH.
abstract: 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 3-6-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)-Ni(2+) complexes, membranes containing PAA/polyethylenimine (PEI)/PAA films bind 93 mg of histidine(6)-tagged (His-tagged) ubiquitin per cm(3) 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 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/cm(3). Preliminary studies suggest that these membranes are stable under depyrogenation conditions (1 M NaOH).
authors.affiliation: Department of Chemistry,  Michigan State University , East Lansing, Michigan 48824, United States.
authors.forename: Somnath
authors.initials: S
authors.lastname: Bhattacharjee
authors.forename: Jinlan
authors.initials: J
authors.lastname: Dong
authors.forename: Yiding
authors.initials: Y
authors.lastname: Ma
authors.forename: Stacy
authors.initials: S
authors.lastname: Hovde
authors.forename: James H
authors.initials: JH
authors.lastname: Geiger
authors.forename: Gregory L
authors.initials: GL
authors.lastname: Baker
authors.forename: Merlin L
authors.initials: ML
authors.lastname: Bruening
mesh_terms: D000180:Acrylic Resins / Q000737:chemistry*; D000327:Adsorption; D000818:Animals; D002417:Cattle; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006900:Hydroxylation; D008567:Membranes, Artificial; D009571:Nitrilotriacetic Acid / Q000031:analogs & derivatives / Q000737:chemistry; D009757:Nylons / Q000737:chemistry; D009942:Organometallic Compounds / Q000737:chemistry; D010539:Permeability; D011073:Polyamines / Q000737:chemistry; D011094:Polyethyleneimine / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*
chemical_list: D000180:Acrylic Resins; D008567:Membranes, Artificial; D009757:Nylons; D009942:Organometallic Compounds; D011073:Polyamines; D011506:Proteins; C063994:polyallylamine; C088321:nickel nitrilotriacetic acid; C006903:carbopol 940; D011094:Polyethyleneimine; D009571:Nitrilotriacetic Acid