Involvement of non‐conserved residues important for PGE 2 binding to the constrained EP3 eLP 2 using NMR and site‐directed mutagenesis

A peptide constrained to a conformation of second extracellular loop of human prostaglandin‐E 2 (PGE 2 ) receptor subtype3 (hEP3) was synthesized. The contacts between the peptide residues at S211 and R214, and PGE 2 were first identified by NMR spectroscopy. The results were used as a guide for site‐directed mutagenesis of the hEP3 protein. The S211L and R214L mutants expressed in HEK293 cells lost binding to [ 3 H]PGE 2 . This study found that the non‐conserved S211 and R214 of the hEP3 are involved in PGE 2 recognition, and implied that the corresponding residues in other subtype receptors could be important to distinguish the different configurations of PGE 2 ligand recognition sites.