Active human ferritin H/L-hybrid and sequence effect on folding efficiency in Escherichia coli

Jeewon Lee, Sung Woo Kim, Yang Hoon Kim, Ji Young Ahn

    Research output: Contribution to journalArticlepeer-review

    17 Citations (Scopus)

    Abstract

    In Escherichia coli, the recombinant human L-chain ferritin was synthesized in the form of inclusion bodies under the control of T7 promoter system. We developed a recombinant ferritin H/L-hybrid by a direct gene fusion between H- and L-chain subunits. Surprisingly, the presence of heavy-chain polypeptide at the amino terminus of light chain significantly increased the cytoplasmic solubility of the recombinant ferritin hybrid, i.e., more than 80% of synthesized ferritin hybrid was soluble in intracellular region regardless of the changes in cell growth and gene expression conditions such as type of inducer, growth media, culture scale, etc. The soluble ferritin H/L-hybrid was biologically active with the iron storage capacity (295 mol Fe+3 per mol H/L-hybrid) equivalent to ferritin standard. Different types of hybrid mutants were also developed using various H-chain derivatives. Comparison of the intracellular solubilities of the synthesized hybrid mutants showed that the N-terminus four helices of heavy subunit were of crucial importance in maintaining the high solubility in E. coli cytoplasm. Consequently, the increased solubility of the ferritin hybrid seems to be related to such H-chain sequence that forms ferroxidase center and promotes effective intra-molecular interaction with L-chain domain of H/L-hybrid for enhancing the folding efficiency.

    Original languageEnglish
    Pages (from-to)225-229
    Number of pages5
    JournalBiochemical and biophysical research communications
    Volume298
    Issue number2
    DOIs
    Publication statusPublished - 2002

    Bibliographical note

    Funding Information:
    The work for stable synthesis of recombinant ferritin H/L-hybrid with bioactivity was supported by Bioprogen Co. The research for correlating H-chain sequence with intracellular solubility of various ferritin hybrids was supported by a Korea University grant.

    Keywords

    • Cytoplasmic solubility
    • Ferritin H/L-hybrid
    • Folding efficiency
    • Intra-molecular interaction
    • Iron storage capacity

    ASJC Scopus subject areas

    • Biophysics
    • Biochemistry
    • Molecular Biology
    • Cell Biology

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