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J Biol Chem

Title:		"Identification of functionally important residues of the silkmoth pheromone biosynthesis-activating neuropeptide receptor, an insect ortholog of the vertebrate neuromedin U receptor"
Author(s):		Kawai T; Katayama Y; Guo L; Liu D; Suzuki T; Hayakawa K; Lee JM; Nagamine T; Hull JJ; Matsumoto S; Nagasawa H; Tanokura M; Nagata K;
Address:		"From the Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. the Molecular Entomology Laboratory, RIKEN Advanced Science Institute, Wako, Saitama 351-0198, Japan, and. the United States Department of Agriculture-Arid Land Agricultural Research Center, Maricopa, Arizona 85138. From the Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan, amtanok@mail.ecc.u-tokyo.ac.jp. From the Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan, aknagata@mail.ecc.u-tokyo.ac.jp"
Journal Title:		J Biol Chem
Year:		2014
Volume:		20140520
Issue:		27
Page Number:		19150 - 19163
DOI:		10.1074/jbc.M113.488999
ISSN/ISBN:		1083-351X (Electronic) 0021-9258 (Print) 0021-9258 (Linking)
Abstract:		"The biosynthesis of sex pheromone components in many lepidopteran insects is regulated by the interaction between pheromone biosynthesis-activating neuropeptide (PBAN) and the PBAN receptor (PBANR), a class A G-protein-coupled receptor. To identify functionally important amino acid residues in the silkmoth PBANR, a series of 27 alanine substitutions was generated using a PBANR chimera C-terminally fused with enhanced GFP. The PBANR mutants were expressed in Sf9 insect cells, and their ability to bind and be activated by a core PBAN fragment (C10PBAN(R2K)) was monitored. Among the 27 mutants, 23 localized to the cell surface of transfected Sf9 cells, whereas the other four remained intracellular. Reduced binding relative to wild type was observed with 17 mutants, and decreased Ca(2+) mobilization responses were observed with 12 mutants. Ala substitution of Glu-95, Glu-120, Asn-124, Val-195, Phe-276, Trp-280, Phe-283, Arg-287, Tyr-307, Thr-311, and Phe-319 affected both binding and Ca(2+) mobilization. The most pronounced effects were observed with the E120A mutation. A molecular model of PBANR indicated that the functionally important PBANR residues map to the 2nd, 3rd, 6th, and 7th transmembrane helices, implying that the same general region of class A G-protein-coupled receptors recognizes both peptidic and nonpeptidic ligands. Docking simulations suggest similar ligand-receptor recognition interactions for PBAN-PBANR and the orthologous vertebrate pair, neuromedin U (NMU) and NMU receptor (NMUR). The simulations highlight the importance of two glutamate residues, Glu-95 and Glu-120, in silkmoth PBANR and Glu-117 and Glu-142 in human NMUR1, in the recognition of the most functionally critical region of the ligands, the C-terminal residue and amide"
Keywords:		Amino Acid Sequence Animals Binding Sites *Bombyx Calcium/metabolism *Computational Biology Conserved Sequence Glutamic Acid Humans Intracellular Space/metabolism Ligands Molecular Docking Simulation Molecular Sequence Data Neuropeptides/*chemistry/*metab;
Notes:		"MedlineKawai, Takeshi Katayama, Yukie Guo, Linjun Liu, Desheng Suzuki, Tatsuya Hayakawa, Kou Lee, Jae Min Nagamine, Toshihiro Hull, J Joe Matsumoto, Shogo Nagasawa, Hiromichi Tanokura, Masaru Nagata, Koji eng Research Support, Non-U.S. Gov't 2014/05/23 J Biol Chem. 2014 Jul 4; 289(27):19150-63. doi: 10.1074/jbc.M113.488999. Epub 2014 May 20"