tailieunhanh - Báo cáo khoa học: A profile of the residues in the second extracellular loop that are critical for ligand recognition of human prostacyclin receptor

The residues in the second extracellular loop (eLP2) of the prostanoid receptors, which are important for specific ligand recognition, were previ-ously predicted in our earlier studies of the thromboxane A2 receptor (TP) using a combination of NMR spectroscopy and recombinant protein approaches. | ễFEBS Journal A profile of the residues in the second extracellular loop that are critical for ligand recognition of human prostacyclin receptor Feng Ni Shui-Ping So Vanessa Cervantes and Ke-He Ruan The Department of Pharmacologicaland PharmaceuticalSciences and The Center for ExperimentalTherapeutics and PharmacoInformatics University of Houston TX USA Keywords G-protein-coupling receptor prostacyclin prostacyclin receptor prostaglandin I2 prostanoid receptor Correspondence . Ruan The Center for Experimental Therapeutics and PharmacoInformatics and Department of Pharmacological and PharmaceuticalSciences University of Houston 521 Science Research Building 2 Houston TX 77204-5037 USA Fax 1 713 743 1884 Tel 1 713 743 1771 E-mail khruan@ Present address Fujian Academe of TraditionalChinese Medicine and Pharmacy Fuzhou Fujian China Received 24 August 2007 revised 2 November 2007 accepted 7 November 2007 doi The residues in the second extracellular loop eLP2 of the prostanoid receptors which are important for specific ligand recognition were previously predicted in our earlier studies of the thromboxane A2 receptor TP using a combination of NMR spectroscopy and recombinant protein approaches. To further test this hypothesis another prostanoid receptor the prostacyclin receptor IP which has opposite biological characteristics to that of TP was used as a model for these studies. A set of recombinant human IPs with site-directed mutations at the nonconserved eLP2 residues were constructed using an Ala-scanning approach and then expressed in HEK293 and COS-7 cells. The expression levels of the recombinant receptors were six-fold higher in HEK293 cells than in COS-7 cells. The residues important for ligand recognition and binding within the N-terminal segment G159 Q162 and C165 and the C-terminal segment L172 R173 M174 and P179 of IP eLP2 were identified by mutagenesis analyses. The molecular mechanisms for the specific ligand .