In vitro refolding of recombinant human free secretory component using equilibrium gradient dialysis

Earl Prinsloo, Vaughan Oosthuizen, Koji Muramoto, Ryno J. Naude

    Research output: Contribution to journalArticlepeer-review

    14 Citations (Scopus)

    Abstract

    Human secretory component (SC) is associated with secretory immunoglobulins (IgA and IgM) and serves to protect the immunoglobulin in the harsh mucosal environment. SC is derived from the polymeric immunoglobulin receptor (pIgR) which transports polymeric immunoglobulins across epithelial cells into secretions. In this present study, we describe the first cloning, expression, in vitro refolding and purification of a free form of human secretory component (rSC) containing the five functional ligand binding domains using Escherichia coli BL21 (DE3). Free rSC was refolded from inclusion bodies by equilibrium dialysis after purification by nickel affinity chromatography under denaturing conditions. Refolded rSC was purified by gel filtration chromatography. Surface plasmon resonance and dot blot association analysis have shown that purified rSC binds IgM with a physiological equilibrium dissociation constant (KD) of 4.6 × 10-8 M and shares structural similarity to native SC. This provides an important step in the elucidation of the structure of this immunologically important receptor.

    Original languageEnglish
    Pages (from-to)179-185
    Number of pages7
    JournalProtein Expression and Purification
    Volume47
    Issue number1
    DOIs
    Publication statusPublished - 2006 May

    Keywords

    • Fc receptors
    • Human
    • Immunoglobulins
    • Polymeric immunoglobulin receptor
    • Secretory component

    ASJC Scopus subject areas

    • Biotechnology

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