Purpose: To investigate tetracycline-inducible expression system for producing clinically usable, high-quality liver X receptor ligand-binding domain recombinant protein

Methods: In this study, we have expressed and purified the recombinant liver X receptor β-ligand binding domain proteins in E. coli using a tetracycline inducible system. To allow for biological activities, we subcloned into pPROTet.E HN vector, expressed in E. coli cells under optimized conditions, purified and characterized the recombinant liver X receptor β-ligand-binding domain proteins using fluorescence polarization assay.

Results: The use of pPROTet.E HN vector simplified downstream purification processes, including cleavage and elution thereby increasing the solubility and yield of the protein of interest. There was a 2.3-fold increase in the efficiency of recombinant LXR β-ligand binding domain (LBD) production by optimizing the expression temperature to 15 °C when compared to those induced at 37 ºC during the induction procedures. A typical dose-response curve obtained using increasing concentrations of the purified recombinant LXR β-LBD (197-461) and measuring fluorescence intensity (FI) as an index of fluorescent peptide binding to LBD showed 50 % effective dose (ED₅₀) value of 533 nM. The recombinant LXR β-LBDs were substantially soluble and should be useful for future biological, biophysical and structural analyses of nuclear receptor complexes. This may represent a new approach to high expression of other nuclear receptors and may be useful as well for other classes of heterodimeric protein partners.

Conclusion: These findings indicate that recombinant LXR β-LBD protein is a promising target for the development of molecular ligands with improved therapeutic windows.