An AIL/IL-based liquid/liquid extraction system for the purification of His-tagged proteins

Appl Microbiol Biotechnol. 2014 Jun;98(12):5665-75. doi: 10.1007/s00253-014-5737-0. Epub 2014 Apr 18.

Abstract

A sorbent based on affinity ionic liquid (AIL), triazacyclononane-ionic liquid, was synthesized, characterized, and applied to the extraction of histidine (His)-tagged proteins from aqueous buffer to ionic liquid (IL) phase. The adsorbed His-tagged proteins could be back-extracted from the IL phase to the aqueous buffer with an imidazole solution. The specific binding of His-tagged proteins with AIL/IL could be affected by a few factors including the ionic strength and coordinated metal ions. In the case of His-tagged enhanced green fluorescent protein (EGFP), the maximum binding capacity of Cu(2+)-AIL/IL reached 2.58 μg/μmol under the optimized adsorption conditions. The eluted His-tagged EGFP kept fluorescent and remained active through the purification process. Moreover, a tandem extraction process successively using Cu(2+)-AIL/IL and Zn(2+)-AIL/IL systems was developed, which was proven very efficient to obtain the ultimate protein with a purity of about 90 %. An effective reclamation method for the AIL/IL extraction system was further established. The sorbent could be easily regenerated by removing metal ions with EDTA and the followed reimmobilization of metal ions. Easy handling of the presented M(2+)-AIL/IL system and highly specific ability to absorb His-tagged proteins make it attractive and potentially applicable in biomolecular separation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adsorption
  • Aza Compounds / chemistry
  • Ionic Liquids / chemistry
  • Liquid-Liquid Extraction / methods*
  • Piperidines / chemistry
  • Proteins / chemistry
  • Proteins / isolation & purification*

Substances

  • Aza Compounds
  • Ionic Liquids
  • Piperidines
  • Proteins
  • triazacyclononane