A two-in-one expression construct for biophysical and structural studies of the human pregnane X receptor...

The ligand-binding domain (LBD) of the human nuclear receptor pregnane X receptor (PXR) is known to crystallize in two different crystal forms, P2₁2₁2₁ or P4₃2₁2, depending on the construct and the strategy used for protein production, as well as the presence or absence of the coactivator-derived peptide SRC-1. In order to facilitate biophysical and structural studies, a versatile construct was designed that allows access to both forms. This was achieved by introducing a thrombin cleavage site between the PXR_LBD and the SRC-1 peptide fused to its C-terminus. Here, we describe the expression, purification and crystallization processes of this novel construct and report two new structures of PXR_LBD that were obtained thanks to this strategy.