Engineering novel specificities for ligand-activated transcription in the nuclear hormone receptor RXR.

ABSTRACT

BACKGROUND: The retinoid X receptor (RXR) activates transcription of target genes in response to its natural ligand, 9-cis retinoic acid (9cRA), and a number of RXR-specific synthetic ligands. To discover the potential for engineering nuclear receptors for activation of transcription by novel ligands, we used structure-based mutagenesis to change the ligand specificity of RXR. RESULTS: By making substitutions at only two positions (Phe313 and Leu436) we engineered two new classes of RXR proteins that had altered ligand specificities. The first class exhibits decreased activation by 9cRA and increased activation by synthetic ligands. The second class continues to be activated by 9cRA but no longer responds to synthetic ligands. The magnitude of the change in specificity that can be accomplished is greater than 280-fold. CONCLUSIONS: These results confirm that Phe313 and Leu436 are crucial determinants of ligand specificity for RXR and demonstrate that nuclear receptors are exceptionally promising protein scaffolds for the introduction of novel ligand specificities through structure-based protein engineering.