Display of aggregation-prone ligand binding domain of human PPAR gamma on surface of bacteriophage lambda

Aim: To display the aggregation-prone ligand binding domain (LBD) of the human
peroxisome proliferator-activated receptor gamma (PPARγ) on the surface of
bacteriophages to establish an easy screening assay for the identification of PPARγ
ligands. Methods: Plasmids were constructed for the expression of the PPARγ
LBD as a fusion to the N-terminus of the g3p protein of filamentous phage or the
C-terminus of the capsid protein D (pD) of phage lambda. The fusion proteins
were expressed in E coli and solubility characteristics were compared. Polyclonal
antibodies against the LBD as well as the pD protein were prepared for Western
blot analysis and phage capture assay. Results: The pD-LBD fusion protein was
partially soluble, whereas the LBD-g3p fusion protein was detected only in the
insoluble fraction. The pD-LBD fusion protein was efficiently incorporated in
phage particles. Furthermore, the LBD was shown to be displayed on the surface
of bacteriophage lambda. On average, the pD-LBD fusion protein accounted for
28% of the total pD protein in the lambda head capsid. Conclusion: The hydrophobic
PPARγ LBD was expressed as a soluble form of fusion protein in E coli and
displayed on the surface of bacteriophage lambda when it was fused to the lambda
pD protein. The lambda pD fusion system could be used for improving the solubility
of proteins that tend to form inclusion bodies when expressed in E coli. The
lambda phage particles displaying the LBD of PPARγ may be of great value for the
identification of novel PPARγ ligands.