title: Identification of important residues of insulin-like peptide 5 and its receptor RXFP4 for ligand-receptor interactions.
abstract: Insulin-like peptide 5 (INSL5) is an insulin/relaxin superfamily peptide involved in the regulation of glucose homeostasis by activating its receptor RXFP4, which can also be activated by relaxin-3 in vitro. To determine the interaction mechanism of INSL5 with its receptor RXFP4, we studied their electrostatic interactions using a charge-exchange mutagenesis approach. First, we identified three negatively charged extracellular residues (Glu100, Asp104 and Glu182) in human RXFP4 that were important for receptor activation by wild-type INSL5. Second, we demonstrated that two positively charged B-chain Arg residues (B13Arg and B23Arg) in human INSL5 were involved in receptor binding and activation. Third, we proposed probable electrostatic interactions between INSL5 and RXFP4: the B-chain central B13Arg of INSL5 interacts with both Asp104 and Glu182 of RXFP4, meanwhile the B-chain C-terminal B23Arg of INSL5 interacts with both Glu100 and Asp104 of RXFP4. The present electrostatic interactions between INSL5 and RXFP4 were similar to our previously identified interactions between relaxin-3 and RXFP4, but had subtle differences that might be caused by the different B-chain C-terminal conformations of relaxin-3 and INSL5 because a dipeptide exchange at the B-chain C-terminus significantly decreased the activity of INSL5 and relaxin-3 to receptor RXFP4.
authors.affiliation: Institute of Protein Research, College of Life Sciences and Technology, Tongji University, Shanghai 200092, China; Central Laboratory, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 200120, China.
authors.forename: Xin-Yi
authors.initials: XY
authors.lastname: Wang
authors.affiliation: Institute of Protein Research, College of Life Sciences and Technology, Tongji University, Shanghai 200092, China.
authors.forename: Yu-Qi
authors.initials: YQ
authors.lastname: Guo
authors.affiliation: Institute of Protein Research, College of Life Sciences and Technology, Tongji University, Shanghai 200092, China.
authors.forename: Xiao-Xia
authors.initials: XX
authors.lastname: Shao
authors.affiliation: Central Laboratory, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 200120, China.
authors.forename: Ya-Li
authors.initials: YL
authors.lastname: Liu
authors.affiliation: Central Laboratory, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 200120, China.
authors.forename: Zeng-Guang
authors.initials: ZG
authors.lastname: Xu
authors.affiliation: Institute of Protein Research, College of Life Sciences and Technology, Tongji University, Shanghai 200092, China; Central Laboratory, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 200120, China. Electronic address: zhan-yun.guo@tongji.edu.cn.
authors.forename: Zhan-Yun
authors.initials: ZY
authors.lastname: Guo
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D006801:Humans; D007328:Insulin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000378:metabolism*; D018000:Receptors, Peptide / Q000737:chemistry* / Q000378:metabolism*; D012065:Relaxin / Q000737:chemistry / Q000378:metabolism; D055672:Static Electricity
chemical_list: D007328:Insulin; C084797:Leydig insulin-like protein; D008024:Ligands; D011506:Proteins; C480238:RLN3 protein, human; C480239:RXFP4 protein, human; D043562:Receptors, G-Protein-Coupled; D018000:Receptors, Peptide; D012065:Relaxin
keywords: Electrostatic interaction; INSL5; Ligand–receptor interaction; Mutagenesis; RXFP4