Skip to main content
SpringerLink
Log in

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

  • Methods and protocols
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

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 Cu2+-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 Cu2+-AIL/IL and Zn2+-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 M2+-AIL/IL system and highly specific ability to absorb His-tagged proteins make it attractive and potentially applicable in biomolecular separation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Scheme 1
Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

References

  • Alvarez-Guerra E, Irabien A (2012) Extraction of lactoferrin with hydrophobic ionic liquids. Sep Purif Technol 98:432–440

    Article  CAS  Google Scholar 

  • Ando H, Manabe S, Nakahara Y, Ito Y (2001) Tag-reporter strategy for facile oligosaccharide synthesis on polymer support. J Am Chem Soc 123(16):3848–3849

    Article  CAS  PubMed  Google Scholar 

  • Blesic M, Lopes A, Melo E, Petrovski Z, Plechkova NV, Canongia Lopes JN, Seddon KR, Rebelo LPN (2008) On the self-aggregation and fluorescence quenching aptitude of surfactant ionic liquids. J Phys Chem B 112(29):8645–8650

    Article  CAS  PubMed  Google Scholar 

  • Chaga GS (2001) Twenty-five years of immobilized metal ion affinity chromatography: past, present and future. J Biochem Biophys Methods 49(1):313–334

    Article  CAS  PubMed  Google Scholar 

  • Chaouk H, Hearn MTW (1999) Examination of the protein binding behaviour of immobilised copper (II)-2, 6-diaminomethylpyridine and its application in the immobilised metal ion affinity chromatographic separation of several human serum proteins. J Biochem Biophys Methods 39:161–177

    Google Scholar 

  • Cheung RCF, Wong JH, Ng TB (2012) Immobilized metal ion affinity chromatography: a review on its applications. Appl Microbiol Biotechnol 96(6):1411–1420

    Article  CAS  PubMed  Google Scholar 

  • Donga RA, Khaliq-Uz-Zaman SM, Chan TH, Damha MJ (2006) A novel approach to oligonucleotide synthesis using an imidazolium ion tag as a soluble support. J Org Chem 71(20):7907–7910

    Article  CAS  PubMed  Google Scholar 

  • Du Z, Yu YL, Wang JH (2007) Extraction of proteins from biological fluids by use of an ionic liquid/aqueous two-phase system. Chem Eur J 13(7):2130–2137

    Article  CAS  PubMed  Google Scholar 

  • Graham B, Spiccia L, Hearn MT (2008) Examination of the binding behaviour of several proteins with the immobilized copper(II) complexes of o-, m- and p-xylylene bridged bis(1,4,7-triazacyclononane) macrocycles. J Chromatogr A 1194(1):30–37

    Article  CAS  PubMed  Google Scholar 

  • Gutierrez R, Martín del Valle E, Galan M (2007) Immobilized metal ion affinity chromatography: status and trends. Sep Purif Rev 36(1):71–111

    Article  CAS  Google Scholar 

  • He X, Chan TH (2006) Ionic-tag-assisted oligosaccharide synthesis. Synthesis 2006(10):1645–1651

    Article  Google Scholar 

  • He X, Chan TH (2007) Structurally defined imidazolium-type ionic oligomers as soluble/solid support for peptide synthesis. Org Lett 9(14):2681–2684

    Article  CAS  PubMed  Google Scholar 

  • Huang JY, Lei M, Wang YG (2006) A novel and efficient ionic liquid supported synthesis of oligosaccharides. Tetrahedron Lett 47(18):3047–3050

    Article  CAS  Google Scholar 

  • Huang JY, Li A, Li JR (2011) An efficient approach for the synthesis of oligosaccharides using ionic liquid supported glycosylation. Carbohydr Polym 83(1):297–302

    Article  CAS  Google Scholar 

  • Janzen R, Unger K, Müller W, Hearn M (1990) Adsorption of proteins on porous and non-porous poly(ethyleneimine) and tentacle-type anion exchangers. J Chromatogr A 522:77–93

    Article  CAS  Google Scholar 

  • Jiang W, Prescott M, Devenish RJ, Spiccia L, Hearn MTW (2009) Separation of hexahistidine fusion proteins with immobilized metal ion affinity chromatographic (IMAC) sorbents derived from MN+-tacn and its derivatives. Biotechnol Bioeng 103(4):747–756

    Article  CAS  PubMed  Google Scholar 

  • Kayitmazer AB, Quinn B, Kimura K, Ryan GL, Tate AJ, Pink DA, Dubin PL (2010) Protein specificity of charged sequences in polyanions and heparins. Biomacromolecules 11(12):3325–3331

    Google Scholar 

  • Kohno Y, Saita S, Murata K, Nakamura N, Ohno H (2011) Extraction of proteins with temperature sensitive and reversible phase change of ionic liquid/water mixture. Polym Chem UK 2(4):862–867

    Article  CAS  Google Scholar 

  • Lan Q, Bassi AS, Zhu JX, Margaritis A (2001) A modified Langmuir model for the prediction of the effects of ionic strength on the equilibrium characteristics of protein adsorption onto ion exchange/affinity adsorbents. Chem Eng J 81(1–3):179–186

    Article  CAS  Google Scholar 

  • Meng H, Chen XW, Wang JH (2011) One-pot synthesis of N, N-bis[2-methylbutyl] imidazolium hexafluorophosphate-TiO2 nanocomposites and application for protein isolation. J Mater Chem 21(38):14857–14863

    Article  CAS  Google Scholar 

  • Miao WS, Chan TH (2005) Ionic-liquid-supported peptide synthesis demonstrated by the synthesis of Leu(5)-enkephalin. J Org Chem 70(8):3251–3255

    Article  CAS  PubMed  Google Scholar 

  • Monier M, Abdel-Latif DA (2013) Modification and characterization of PET fibers for fast removal of Hg(II), Cu(II) and Co(II) metal ions from aqueous solutions. J Hazard Mater 250:122–130

    Article  PubMed  Google Scholar 

  • Moniruzzaman M, Kamiya N, Goto M (2010) Activation and stabilization of enzymes in ionic liquids. Org Biomol Chem 8(13):2887–2899

    Article  CAS  PubMed  Google Scholar 

  • Mooney JT, Fredericks D, Hearn MT (2011) Use of phage display methods to identify heptapeptide sequences for use as affinity purification ‘tags’ with novel chelating ligands in immobilized metal ion affinity chromatography. J Chromatogr A 1218(1):92–99

    Article  CAS  PubMed  Google Scholar 

  • Oppermann S, Stein F, Kragl U (2011) Ionic liquids for two-phase systems and their application for purification, extraction and biocatalysis. Appl Microbiol Biotechnol 89(3):493–499

    Article  CAS  PubMed  Google Scholar 

  • Pathak AK, Yerneni CK, Young Z, Pathak V (2008) Oligomannan synthesis using ionic liquid supported glycosylation. Org Lett 10(1):145–148

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Pei Y, Wang J, Wu K, Xuan X, Lu X (2009) Ionic liquid-based aqueous two-phase extraction of selected proteins. Sep Purif Technol 64(3):288–295

    Article  CAS  Google Scholar 

  • Porath J, Carlsson J, Olsson I, Belfrage G (1975) Metal chelate affinity chromatography, a new approach to protein fractionation. Nature 258:598–599

    Article  CAS  PubMed  Google Scholar 

  • Sharma S, Agarwal GP (2001) Interactions of proteins with immobilized metal ions: role of ionic strength and pH. J Colloid Interface Sci 243(1):61–72

    Article  CAS  Google Scholar 

  • Shim Y, Kim HJ, Jung Y (2012) Solvation of a small metal-binding peptide in room-temperature ionic liquids. Bull Korean Chem Soc 33(11):3601–3606

    Article  CAS  Google Scholar 

  • Shimojo K, Kamiya N, Tani F, Naganawa H, Naruta Y, Goto M (2006a) Extractive solubilization, structural change, and functional conversion of cytochrome c in ionic liquids via crown ether complexation. Anal Chem 78(22):7735–7742

    Article  CAS  PubMed  Google Scholar 

  • Shimojo K, Nakashima K, Kamiya N, Goto M (2006b) Crown ether-mediated extraction and functional conversion of cytochrome c in ionic liquids. Biomacromolecules 7(1):2–5

    Article  CAS  PubMed  Google Scholar 

  • Stepanenko OV, Verkhusha VV, Kazakov VI, Shavlovsky MM, Kuznetsova IM, Uversky VN, Turoverov KK (2004) Comparative studies on the structure and stability of fluorescent proteins EGFP, zFP506, mRFP1, “dimer2”, and DsRed1. Biochemistry 43(47):14913–14923

    Article  CAS  PubMed  Google Scholar 

  • Tang S, Baker GA, Zhao H (2012) Ether and alcohol functionalized task-specific ionic liquids: attractive properties and applications. Chem Soc Rev 41(10):4030–4066

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Tseng MC, Tseng MJ, Chu YH (2009) Affinity ionic liquid. Chem Commun 48:7503–7505

    Article  Google Scholar 

  • Ventura SPM, de Barros RLF, de Pinho Barbosa JM, Soares CMF, Lima ÁS, Coutinho JAP (2012) Production and purification of an extracellular lipolytic enzyme using ionic liquid-based aqueous two-phase systems. Green Chem 14(3):734–740

    Article  CAS  Google Scholar 

  • Yerneni CK, Pathak V, Pathak AK (2009) Imidazolium cation supported solution-phase assembly of homolinear alpha(1→6)-linked octamannoside: an efficient alternate approach for oligosaccharide synthesis. J Org Chem 74(16):6307–6310

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Yuan S, Deng Q, Fang G, Pan M, Zhai X, Wang S (2012) A novel ionic liquid polymer material with high binding capacity for proteins. J Mater Chem 22(9):3965–3972

    Article  CAS  Google Scholar 

  • Zachariou M (2004) Immobilized metal ion affinity chromatography of proteins. In: HPLC of peptides and proteins: methods and protocols. Springer, New York, pp 89–102

  • Zachariou M, Hearn MT (1996) Application of immobilized metal ion chelate complexes as pseudocation exchange adsorbents for protein separation. Biochemistry 35(1):202–211

    Article  CAS  PubMed  Google Scholar 

  • Zafarani-Moattar MT, Hamzehzadeh S (2011) Partitioning of amino acids in the aqueous biphasic system containing the water-miscible ionic liquid 1-butyl-3-methylimidazolium bromide and the water-structuring salt potassium citrate. Biotechnol Prog 27(4):986–997

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This study was supported by the Technology Planning Project of the Zhejiang Province of China (2011C12031), the National Science Foundation of China (21102129), and the Opening Foundation of Zhejiang Provincial Top Key Discipline of New Materials and Process Engineering (20121127, 20121128).

Author information

Authors and Affiliations

  1. Key Laboratory of Fruits and Vegetables Postharvest and Processing Technology Research of Zhejiang Province, College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People’s Republic of China

    Weiyuan Xu, Guangwei Ren, Hujun Xie & Jianying Huang

  2. College of Material, Chemistry and Chemical Engineering, Hangzhou Normal University, Hangzhou, 310036, People’s Republic of China

    Shijun Li

  3. College of Chemical Engineering and Materials Science, Zhejiang University of Technology, Hangzhou, 310014, People’s Republic of China

    Huazhen Cao

Authors
  1. Weiyuan Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Huazhen Cao
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Guangwei Ren
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hujun Xie
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jianying Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Shijun Li
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Jianying Huang or Shijun Li.

Electronic supplementary material

Below is the link to the electronic supplementary material.

ESM 1

(PDF 221 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Xu, W., Cao, H., Ren, G. et al. An AIL/IL-based liquid/liquid extraction system for the purification of His-tagged proteins. Appl Microbiol Biotechnol 98, 5665–5675 (2014). https://doi.org/10.1007/s00253-014-5737-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-014-5737-0

Keywords

Search

Navigation