Affinity partitioning of protein a using a magnetic aqueous two-phase system

Motoshi Suzuki, Masamichi Kamihira, Tomoki Shiraishi, Hiroshi Takeuchi, Takeshi Kobayashi

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

An affinity extraction method combining an aqueous two-phase system and affinity ligand-bound fine magnetic particles is proposed as a novel protein separation technique. Polymer-coated magnetic particles, Eudragit-Mag, partitioned to the upper phase stably in three types of PEG-based aqueous two-phase systems. The phase separation time was shortened by the additin of the magnetic particles, and further reduced by applying a magnetic field. Human IgG was immobilized to Eudragit-Mag for the separation of staphylococcal protein. A. The partition coefficient of protein A increased up to 35-fold when IgG-bound Eudragit-Mag was added to a PEG-phosphate aqueous two-phase system. The IgG-bound Eudragit-Mag was applied to the purification of protein A produced by recombinant Escherichia coli. In the separation, 90% of protein A added was selectively adsorbed to the affinity ligand, where the content of protein A was 89%. By using 3.5 M KSCN solution as an eluant, 39% of protein A was recovered with 49% of purity from a crude extract. In addition to the intrinsic advantages of aqueous two-phase extraction, the newly developed separation technique has advantages in the selective partitioning of the target protein, a shortening of phase separation time, and easy recovery and reuse of the ligand.

Original languageEnglish
Pages (from-to)78-84
Number of pages7
JournalJournal of Fermentation and Bioengineering
Volume80
Issue number1
DOIs
Publication statusPublished - 1995
Externally publishedYes

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Staphylococcal Protein A
Proteins
Immunoglobulin G
Ligands
Phase separation
Polyethylene glycols
Magnetic Fields
Complex Mixtures
Escherichia coli
Purification
Polymers
Phosphates
Magnetic fields
Recovery
methylmethacrylate-methacrylic acid copolymer

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Applied Microbiology and Biotechnology

Cite this

Affinity partitioning of protein a using a magnetic aqueous two-phase system. / Suzuki, Motoshi; Kamihira, Masamichi; Shiraishi, Tomoki; Takeuchi, Hiroshi; Kobayashi, Takeshi.

In: Journal of Fermentation and Bioengineering, Vol. 80, No. 1, 1995, p. 78-84.

Research output: Contribution to journalArticle

Suzuki, Motoshi ; Kamihira, Masamichi ; Shiraishi, Tomoki ; Takeuchi, Hiroshi ; Kobayashi, Takeshi. / Affinity partitioning of protein a using a magnetic aqueous two-phase system. In: Journal of Fermentation and Bioengineering. 1995 ; Vol. 80, No. 1. pp. 78-84.
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