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The quiescin sulfhydryl oxidase (hQSOX1b) tunes the expression of resistin-like molecule alpha (RELM-α or mFIZZ1) in a wheat germ cell-free extract.
- Author(s): Gad, Wael;
- Nair, Meera G;
- Van Belle, Karolien;
- Wahni, Khadija;
- De Greve, Henri;
- Van Ginderachter, Jo A;
- Vandenbussche, Guy;
- Endo, Yaeta;
- Artis, David;
- Messens, Joris
- et al.
Published Web Locationhttps://doi.org/10.1371/journal.pone.0055621
BackgroundAlthough disulfide bond formation in proteins is one of the most common types of post-translational modifications, the production of recombinant disulfide-rich proteins remains a challenge. The most popular host for recombinant protein production is Escherichia coli, but disulfide-rich proteins are here often misfolded, degraded, or found in inclusion bodies.
Methodology/principal findingsWe optimize an in vitro wheat germ translation system for the expression of an immunological important eukaryotic protein that has to form five disulfide bonds, resistin-like alpha (mFIZZ1). Expression in combination with human quiescin sulfhydryl oxidase (hQSOX1b), the disulfide bond-forming enzyme of the endoplasmic reticulum, results in soluble, intramolecular disulfide bonded, monomeric, and biological active protein. The mFIZZ1 protein clearly suppresses the production of the cytokines IL-5 and IL-13 in mouse splenocytes cultured under Th2 permissive conditions.
Conclusion/significanceThe quiescin sulfhydryl oxidase hQSOX1b seems to function as a chaperone and oxidase during the oxidative folding. This example for mFIZZ1 should encourage the design of an appropriate thiol/disulfide oxidoreductase-tuned cell free expression system for other challenging disulfide rich proteins.
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