title: Identification by site-directed mutagenesis of amino acids contributing to ligand-binding specificity or signal transduction properties of the human FP prostanoid receptor.
abstract: Prostanoid receptors belong to the class of heptahelical plasma membrane receptors. For the five prostanoids, eight receptor subtypes have been identified. They display an overall sequence similarity of roughly 30%. Based on sequence comparison, single amino acids in different subtypes of different species have previously been identified by site-directed mutagenesis or in hybrid receptors that appear to be essential for ligand binding or G-protein coupling. Based on this information, a series of mutants of the human FP receptor was generated and characterized in ligand-binding and second-messenger-formation studies. It was found that mutation of His-81 to Ala in transmembrane domain 2 and of Arg-291 to Leu in transmembrane domain 7, which are putative interaction partners for the prostanoid's carboxyl group, abolished ligand binding. Mutants in which Ser-263 in transmembrane domain 6 or Asp-300 in transmembrane domain 7 had been replaced by Ala or Gln, respectively, no longer discriminated between prostaglandins PGF(2alpha) and PGD(2). Thus distortion of the topology of transmembrane domains 6 and 7 appears to interfere with the cyclopentane ring selectivity of the receptor. PGF(2alpha)-induced inositol formation was strongly reduced in the mutant Asp-300Gln, inferring a role for this residue in agonist-induced G-protein activation.
authors.affiliation: Universität Potsdam, Institut für Ernährungswissenschaft, Lehrstuhl Biochemie der Ernährung, Arthur-Scheunert-Allee 114-116, 14558 Bergholz-Rehbrücke, D-14558 Bergholz-Rehbrücke, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Neuschäfer-Rube
authors.forename: Eva
authors.initials: E
authors.lastname: Engemaier
authors.forename: Sina
authors.initials: S
authors.lastname: Koch
authors.forename: Ulrike
authors.initials: U
authors.lastname: Böer
authors.forename: Gerhard P
authors.initials: GP
authors.lastname: Püschel
mesh_terms: D019943:Amino Acid Substitution; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D015237:Dinoprost / Q000378:metabolism; D005260:Female; D006801:Humans; D007700:Kinetics; D008024:Ligands; D016297:Mutagenesis, Site-Directed; D010920:Placenta / Q000378:metabolism; D011247:Pregnancy; D011982:Receptors, Prostaglandin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D017931:DNA Primers; D008024:Ligands; D011982:Receptors, Prostaglandin; D011994:Recombinant Proteins; C034671:prostaglandin F2alpha receptor; D015237:Dinoprost