The Relaxin Receptor (RXFP1) Utilizes Hydrophobic Moieties on a Signaling Surface of Its N-terminal Low Density Lipoprotein Class A Module to Mediate Receptor Activation

Abstract

Background: Activation of the relaxin receptor RXFP1 is driven by the LDLa module at the RXFP1 N terminus. Results: LDLa residues Leu-7, Tyr-9, and Lys-17 all contribute to receptor activation via interactions involving their hydrophobic side chains. Conclusion: These interactions induce the active receptor conformation, suggesting a novel mode of GPCR activation. Significance: This novel mechanism of GPCR activation may lead to new drug development.

Investigation of Interactions at the Extracellular Loops of the Relaxin Family Peptide Receptor 1 (RXFP1)

“…Nevertheless, small chemical shift differences were localized to the C-terminal region of the LDLa module, implying that this region interacts with EL1/EL2 of RXFP1. In our previous studies using site-directed mutagenesis and both knock-in and -out functional analyses, we showed that a hydrophobic surface in the N-terminal region of the RXFP1 LDLa comprising Leu 7 , Tyr 9 , and Lys 17 were important for receptor activity (9,10). Importantly, the knock-out mutagenesis experiments (10) could not abolish activation, suggesting that there were critical residues that we had not revealed.…”

“…action demonstrated the possibility of the LDLa module also having access to the TM domain of receptors (44). Functional characterization of LDLa-less, chimeric, and mutant receptors has demonstrated a critical functional role for the LDLa module (10,24). Because of the inability of concentrating EL1/EL2-GB1 beyond ϳ200 M, 15 N-LDLa was titrated to only equimolar with EL1/EL2-GB1.…”

“…Its removal does not affect relaxin binding; however, the receptor does not signal, suggesting that relaxin binding alone is not sufficient to initiate signaling. Mutagenesis studies of the LDLa reveal that residues within the N-terminal region of the module are involved in receptor activation highlighting the possibility that the LDLa module is able to make activating contacts with the TM domain (9,10). Molecular characterization of relaxin and LDLa interactions at the ELs of RXFP1 has been limited because of the inherent difficulties in studying membrane-associated GPCRs.…”

International Union of Basic and Clinical Pharmacology. XCV. Recent Advances in the Understanding of the Pharmacology and Biological Roles of Relaxin Family Peptide Receptors 1–4, the Receptors for Relaxin Family Peptides

“…Further evidence for a specific interaction between the LDLa module and RXFP1 that drives receptor activation comes from studies on chimeric and mutant RXFP1 receptors (Hopkins et al, 2007;Kong et al, 2013). A chimera where the RXFP1 LDLa module is swapped Fig.…”

“…Relaxin binds with high affinity to the LRR region of RXFP1 followed by a lower affinity interaction with ECL2 (Sudo et al, 2003;Halls et al, 2005a,b). This triggers an interaction involving the LDLa module that is essential for signaling Kern et al, 2007;Kong et al, 2013). Mutations of the calcium binding asparagine within the LDLa module (D58E), produce a receptor where the cAMP response to relaxin is completely abolished but that to ML290 (compound 8) is unaffected .…”