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Updated: Jul 11, 2026
Real-time Analyses of Retinol Transport by the Membrane Receptor of Plasma Retinol Binding Protein
Published on: January 28, 2013
Recombinant human retinol-binding protein refolding, native disulfide formation, and characterization
Y Xie1, H A Lashuel, G J Miroy
1Department of Chemistry and the Skaggs Institute of Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, MB12, La Jolla, California, 92037, USA.
Researchers developed a method to refold human retinol-binding protein (RBP) from inclusion bodies in E. coli. This efficient process yields pure RBP, crucial for vitamin A transport and cellular utilization.
Area of Science:
- Biochemistry
- Molecular Biology
- Protein Chemistry
Background:
- Human retinol-binding protein (RBP) is vital for retinol transport.
- Overexpression of RBP in E. coli leads to insoluble inclusion bodies.
Purpose of the Study:
- To develop an efficient method for refolding aggregated RBP.
- To obtain pure, functional RBP for further research.
Main Methods:
- Solubilization of RBP inclusion bodies using GdmCl and DTT.
- Redox refolding in the presence of vitamin A using a cysteine/cystine buffer.
- Purification via ion exchange chromatography (HPLC).
Main Results:
- Refolded RBP was purified to homogeneity, confirmed by SDS-PAGE and MS.
- The refolded RBP demonstrated native structure by binding vitamin A and transthyretin.
- An overall yield of 50-60% (73 mg/L culture) was achieved.
Conclusions:
- A robust protocol for reconstituting functional RBP from E. coli inclusion bodies was established.
- This method provides a high yield of pure RBP, facilitating studies on its biological roles.
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