title: A negatively charged transmembrane aspartate residue controls activation of the relaxin-3 receptor RXFP3.
abstract: Relaxin-3 is an insulin/relaxin superfamily neuropeptide involved in the regulation of food intake and stress response via activation of its cognate receptor RXFP3, an A-class G protein-coupled receptor (GPCR). In recent studies, a highly conserved ExxxD motif essential for binding of relaxin-3 has been identified at extracellular end of the second transmembrane domain (TMD2) of RXFP3. For most of the A-class GPCRs, a highly conserved negatively charged Asp residue (Asp(2.50) using Ballesteros-Weinstein numbering and Asp128 in human RXFP3) is present at the middle of TMD2. To elucidate function of the conserved transmembrane Asp128, in the present work we replaced it with other residues and the resultant RXFP3 mutants all retained quite high ligand-binding potency, but their activation and agonist-induced internalization were abolished or drastically decreased. Thus, the negatively charged transmembrane Asp128 controlled transduction of agonist-binding information from the extracellular region to the intracellular region through maintaining RXFP3 in a metastable state for efficient conformational change induced by binding of an agonist.
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China.
authors.forename: Xiao-Xia
authors.initials: XX
authors.lastname: Shao
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China.
authors.forename: Meng-Jun
authors.initials: MJ
authors.lastname: Hu
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China.
authors.forename: Ya-Li
authors.initials: YL
authors.lastname: Liu
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China.
authors.forename: Zeng-Guang
authors.initials: ZG
authors.lastname: Xu
authors.affiliation: Research Center for Translational Medicine, College of Life Sciences and Technology, Tongji University, Shanghai, China. Electronic address: zhan-yun.guo@tongji.edu.cn.
authors.forename: Zhan-Yun
authors.initials: ZY
authors.lastname: Guo
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D005456:Fluorescent Dyes / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000737:chemistry; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011485:Protein Binding; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D012065:Relaxin / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D008024:Ligands; C480238:RLN3 protein, human; C410092:RXFP3 protein, human; D043562:Receptors, G-Protein-Coupled; D049452:Green Fluorescent Proteins; D001224:Aspartic Acid; D012065:Relaxin
keywords: GPCR; Ligand binding; RXFP3; Receptor activation; Relaxin-3