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Refolding of fusion ferritin by gel filtration chromatography (GFC)

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Abstract

Fusion ferritin (heavy chain ferritin, FH+light chain ferritin, FL), an iron-binding protein, was primarily purified from recombinantEscherichia coli by two-step sonications with urea [1]. Unfolded ferritin was refolded by gel filtration chromatography (GFC) with refolding enhancer, where 50 mM Na-phosphate (pH 7.4) buffer containing additives such as Tween 20, PEG, andl-arginine was used. Ferritin is a multimeric protein that contains approximately 20 monomeric units for full activity. Fusion ferritin was expressed in the form of inclussion bodies (Ibs). The IBs were initially solubilized in 4 M urea denaturant. The refolding process was then performed by decreasing the urea concentration on the GFC column to form protein multimers. The combination of the buffer-exchange effect of GFC and the refolding enhancers in refolding buffer resulted in an efficient route for producing properly folded fusion ferritin.

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References

  1. Huh, Y. S. and I. H. Kim (2003) Purification fusion ferritin from recombinantE. coli using two-step sonications.Biotechnol. Lett. 25: 993–996.

    Article  CAS  Google Scholar 

  2. Shin, H. C. (2001) Protein folding, misfolding, and refolding of therapeutic proteins.Biotechnol. Bioprocess Eng. 6: 237–243.

    Article  CAS  Google Scholar 

  3. Lilie, H., E. Schwarz, and R. Rudolph (1998) Advances in refolding of proteins produced inE. coli.Curr. Opin. Biotechnol. 9: 497–501.

    Article  CAS  Google Scholar 

  4. Fahey, E. M., J. B. Chaudhuri, and P. Binding (2000) Refolding and purification of a urokinase plasminogen activator fragment by chromatography.J. Chromatogr. B 737: 225–235.

    Article  CAS  Google Scholar 

  5. Guise A. D. and J. B. Chaudhuri (2001) Initial protein concentration and residual denaturant concentration strongly affect the batch refolding of hen egg white lysozyme.Biotechnol. Bioprocess Eng. 6: 410–418.

    Article  CAS  Google Scholar 

  6. Geng, X. D. and X. Q. Chang (1992) High-performance hydrophobic interaction chromatography as a tool for protein refolding.J. Chromatogr. A 599: 185–194.

    Article  CAS  Google Scholar 

  7. Guan Y.-X., H.-X. Pan, Y.-G. Gao, S.-J. Yao, and M. G. Cho (2005) Refolding and purification of recombinant human interferon-8 expressed as inclusion bodies inE. coli using size-exclusion chromatography.Biotechnol. Bioprocess Eng. 10: 122–127.

    Article  CAS  Google Scholar 

  8. Lee, G., Y. H. Y. Song, H. T. Chung, M. J. Youn, and S. I. Jang (1991) Differential effects of ferritin on the humoral and cellular immune responses in the mouse.Molecular Cells. 1: 169–175.

    Google Scholar 

  9. Kim, K. S., S. R. Chang, and Y. T. Kim (1995) Purification and characterization of recombinant tadpole H-chain ferritin inE. coli.J. Biochem. Mol. Biol. 28: 238–242.

    Google Scholar 

  10. Lee, J., H. Y. Seo, E. S. Jeon, O. S. Park, K. M. Lee, C. U. Park, and K. S. Kim (2001) Cooperative activity of subunits of human ferritin heteropolymers in.E. coli.J. Biochem. Mol. Biol. 34: 365–370.

    CAS  Google Scholar 

  11. Kim, K. S. (1998) Influence of site directed mutagenesis on protein assembly and solubility of tadpole H-chain ferritin.Biotechnol. Bioprocess Eng. 3: 67–70.

    Article  Google Scholar 

  12. Bradford, M. M. (1976) A rapid and sensitive method for the quantization of protein utilizing the principles of protein-dye binding.Anal. Biochem. 72: 248–254.

    Article  CAS  Google Scholar 

  13. Laemmli, U. K. (1970) Detection of structural proteins during the assembly of the head of Bacteriophage T4.Nature 227: 680–685.

    Article  CAS  Google Scholar 

  14. Kim, S. W., Y. H. Kim, and J. W. Lee (2001) Thermal stability of human ferritin: concentration dependence and enhanced stability of an N-terminal fusion mutant.Biochem. Biophys. Res. Commun. 289: 125–129.

    Article  CAS  Google Scholar 

  15. Gao, Y. G., Y. X. Guan, S. J. Yao, and M. G. Cho (2002) Refolding of lysozyme at high concentration in batch and fed-batch operation.Korean J. Chem. Eng. 19: 871–875.

    Article  CAS  Google Scholar 

  16. Lee, J.-H., N.-G. Loc, T.-H. Kwon, and M.-S. Yang (2004) Partitioning of recombinant human granulocyte macrophage colony stimulating factor from plant suspension culture in PEG/sodium phosphate aqueous two-phase systems,Biotechnol. Bioprocess Eng. 9: 12–16.

    Article  CAS  Google Scholar 

  17. Hong, S. P., C. H. Lee, S. K. Kim, H. S. Yun, J. H. Lee, and K. H. Row (2004) Mobile phase compositions for ceramide III by normal phase high performance liquid chromatography.Biotechnol. Bioprocess Eng. 9: 47–51.

    Article  CAS  Google Scholar 

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  1. Department of Chemical Engineering, Chungnam National University, 305-764, Daejeon, Korea

    Hyungwon Kim & In Ho Kim

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  1. Hyungwon Kim
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  2. In Ho Kim
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Correspondence to In Ho Kim.

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Kim, H., Kim, I.H. Refolding of fusion ferritin by gel filtration chromatography (GFC). Biotechnol. Bioprocess Eng. 10, 500–504 (2005). https://doi.org/10.1007/BF02932284

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  • DOI: https://doi.org/10.1007/BF02932284

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