Purpose: To investigate tetracycline-inducible expression system for producing clinically usable, highquality
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°C 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 (ED
50) 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.
