Preparation of a modified crosslinked chitosan/polyvinyl alcohol blended affinity membrane for purification of His‐tagged protein

Abstract

Based on a crosslinked chitosan (CS)/polyvinyl alcohol (PVA) matrix membrane, an immobilized metal ion affinity membrane (IMAM) using Cu 2+ and Ni 2+ ions as affinity ligands was prepared for purification of the His-tagged recombinant protein. The affinity membrane possessed a favorable membrane structure including 1.39 μm average pore size and 0.33 mLÁcm −2 Ás −1 water flux under 0.08 MPa pressure at 25 C. The Cu 2+ and Ni 2+ ions capacities immobilized on the IMAM were 155.6 and 137.3 μmolÁdisk −1 , respectively. The IMAM had an excellent specific affinity to His-tagged protein. About 10-fold purification factor for the model protein was obtained in a batch adsorption, and serine hydroxymethyl transferase could be purified to a single band in sodium dodecyl sulfatepolyacrylamide gel electrophoresis analysis from its crude extract solution with an affinity membrane cartridge by a dynamic purification process. This work provides a promising IMAM for the purification of His-tagged recombinant proteins.

Recent Developments of Liquid Chromatography Stationary Phases for Compound Separation: From Proteins to Small Organic Compounds

“…Complex formation commonly occurs in chromatography where the analytes are proteins [ 1 , 2 , 3 ], DNA [ 4 ], and RNA [ 5 , 6 ]. The stationary phase is modified so that it has a metal-complex binding site that can accept ligands from macromolecules.…”

Recent trends in sorbents for bioaffinity chromatography

“…Likewise, Cu (II), Ni (II), Co (II), and Zn (II) cations are also used to purify proteins containing histidine residues [47][48][49][50]. High selectivity is achieved by the interaction of the immobilized cation with the imidazole ring of the histidine residues in peptides and proteins [47][48][49][50]. IMAC was first proposed by Porath et al using an affinity sorbent, which was prepared by the immobilization of metal cation (Ni(II) and Cu(II)) on agarose using IDA as a chelating ligand, for the purification of histidine-containing proteins [51].…”

“…Likewise, Cu (II), Ni (II), Co (II), and Zn (II) cations are also used to purify proteins containing histidine residues [47–50]. High selectivity is achieved by the interaction of the immobilized cation with the imidazole ring of the histidine residues in peptides and proteins [47–50]. IMAC was first proposed by Porath et al.…”

“…In the last decade, there has been an increasing trend in the enrichment of biomarkers with IMAC in various biotechnological and biomedical applications before mass spectrometric analysis for their detection in biological samples [7, 9, 42–44, 47–50, 14,15,51–60]. IMAC has become an important technique in the purification of histidine‐tagged proteins and phosphoproteins in recent years.…”

Polyvinyl alcohol/multi‐walledcarbon nanotubes nanocomposites with ordered macroporous structures prepared byice‐templating

“…Polyvinyl alcohol (PVA) is a versatile and nontoxic polymer used in tissue engineering, 22,23 adsorption, 24,25 supercapacitors, 26,27 and in the textile and food industries 28 . The preparation of PVA‐based scaffolds has also been widely investigated over the past years due to its technological importance in several applications 29–31 . In this study, we prepared PVA/multi‐walled carbon nanotubes (MWCNTs) macroporous nanocomposites by ice‐templating.…”