Amino acid residues conferring ligand binding properties of prostaglandin I and prostaglandin D receptors. Identification by site-directed mutagenesis.
Using chimeras of the mouse prostaglandin (PG) I receptor (mIP) and the mouse PGD receptor (mDP), we previously revealed that the cyclopentane ring recognition by these receptors is specified by a region from the first to third transmembrane domain of each receptor; recognition by this region of mIP is broad, accommodating the D, E, and I types of cyclopentane rings, whereas that of mDP binds the D type of PGs alone (Kobayashi, T., Kiriyama, M., Hirata, T., Hirata, M., Ushikubi, F., and Narumiya, S. (1997) J. Biol. Chem. 272, 15154-15160). In the present study, we performed a more detailed chimera analysis, and narrowed the domain for the ring recognition to a region from the first transmembrane domain to the first extracellular loop. One chimera with the replacement of the second transmembrane domain and the first extracellular loop of mDP with that of mIP bound only iloprost. The amino acid substitutions in this chimera suggest that Ser(50) in the first transmembrane domain of mIP confers the broad ligand recognition of mIP and that Lys(75) and Leu(83) in the second transmembrane domain of mDP confer the high affinity to PGD(2) and the strict specificity of ligand binding of mDP, respectively.
Kobayashi
T
T
Department of Pharmacology, Faculty of Medicine, Kyoto University, Kyoto 606-8501, Japan.
Ushikubi
F
F
Narumiya
S
S
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D017931:DNA Primers; D007700:Kinetics; D007930:Leucine; D008024:Ligands; D008239:Lysine; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D044006:Receptors, Epoprostenol; D011971:Receptors, Immunologic; D011982:Receptors, Prostaglandin / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012694:Serine
D017931:DNA Primers; D008024:Ligands; D044006:Receptors, Epoprostenol; D011971:Receptors, Immunologic; D011982:Receptors, Prostaglandin; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012694:Serine; D007930:Leucine; D008239:Lysine; C036293:prostaglandin D2 receptor