A Pluronic-coupled metal-chelating ligand for membrane affinity chromatography
Date
2006
Authors
Govender S.
Przybylowicz W.J.
Jacobs E.P.
Bredenkamp M.W.
Kralingen L.v.
Swart P.
Journal Title
Journal ISSN
Volume Title
Publisher
Abstract
A technique for bio-specific affinity chromatography using synthetic nonporous membranes and a new metal-chelating Pluronic surfactant is described. Synthetic polymeric poly(vinyldiene fluoride) membranes were fabricated for use as solid, hydrophobic adsorption membrane matrices. An ethylene diamine tetraacetic acid dianhydride was coupled to the terminal hydroxyl end groups of Pluronic® F108 via a two-step reaction at 40 °C, to create a new metal affinity ligand, Pluronic-N,N-dicarboxymethyl-3,6-diazaoctanedioate (Pluronic-DMDDO). The hydrophobic poly(propylene oxide) moiety of Pluronic allowed non-covalent adsorption of the ligand to the hydrophobic membrane matrix. The protein repellent properties of the hydrophilic poly(ethylene oxide) brush layer of Pluronic, served to preserve the bio-specific activity of the ligand and to increase ligand accessibility. Proton-induced X-ray emission (PIXE) analysis was used to determine the metal binding capacity, stability and surface homogeneity of this immobilised metal affinity membrane system and to generate surface homogeneity maps of the chelated metal ions. The chelate capacity of Pluronic-DMDDO was determined under non-competitive conditions and was of the order (Zn2+ > Ni2+ > Cu2+). An amino terminal hex-histidine-tagged recombinant Escherichia coli pantothenate kinase was used as a test protein. Histidine-tagged pantothenate kinase bound strongly to Pluronic-DMDDO-treated membranes specifically in the presence of Ni2+. Eluted immobilised histidine-tagged proteins retained their biochemical activity and the membranes were capable of being regenerated and re-used. © 2005 Elsevier B.V. All rights reserved.
Description
Keywords
Adsorption, Affinity chromatography, Chelation, Hydrophobicity, Proteins, Surface active agents, Membrane affinity chromatography, Metal-chelating ligands, Pluronic F108, Proton-induced X-ray emission (PIXE), Biological membranes, acid anhydride, ethylenediamine, histidine, hydroxyl group, ligand, metal chelate, metal ion, pantothenate kinase, poloxamer, polymer, Adsorption, Affinity chromatography, Biological membranes, Chelation, Hydrophobicity, Proteins, Surface active agents, adsorption, affinity chromatography, amino terminal sequence, article, binding affinity, biochemistry, Escherichia coli, hydrophilicity, hydrophobicity, membrane, porosity, priority journal, solid, X ray analysis
Citation
Journal of Membrane Science
279
02-Jan
279
02-Jan