posters 5th Asia-Pacific NMR Symposium 2013

Is the linker between the relaxin-binding and activation domains of its receptor important for signaling? (#199)

Ashish Sethi 1 , Emma J. Petrie 1 , Shoni Bruell 2 , Roy C.Z Kong 2 , Ross A.D. Bathgate 2 , Paul R. Gooley 1
  1. Bio21 Institute, University of Melbourne, Parkville, VIC, Australia
  2. Florey Department of Neuroscience and Mental Health, Florey Institute of Neuroscience and Mental Health , Melbourne, VICTORIA, AUSTRALIA

The Relaxin Receptor (RXFP1) is a unique G-protein coupled receptor (GPCR) with a large extracellular domain consisting of Leucine Rich Repeats (LRRs) tethered by a 34 residue linker to a Low Density Lipoprotein Class-A module (LDLa). RXFP1 is activated by the peptide hormone relaxin, initially considered as a reproductive hormone, but has been demonstrated to be an effective treatment for acute heart failure in a recently completed Phase III clinical trial.

Relaxin has an insulin-like chain structure and binds primarily to the LRRs region via specific interactions including glutamate/aspartate residues in the receptor with an arginine cassette on the B-chain of the hormone. It is proposed that once bound to the LRRs, the A- chain of relaxin interacts with the trans-membrane loops of the receptor; however relaxin binding alone does not elicit signaling. The LDLa module was found to be indispensable for receptor signaling as RXFP1 receptor with the LDLa module removed still binds to relaxin but do not signal.

We hypothesize that the 34 residue LRR-LDLa linker is pivotal in creating the active conformation of the receptor that leads to signal activation involving the LDLa module. We have shown that any insertion/deletion of the LRR-LDLa linker residues leads to the loss of receptor signaling via cAMP. Additionally, we have recombinantly expressed the LDLa module and 34 residue linker (LDLa+) and detected relaxin binding in a titration monitored by 1H-15N Heteronuclear Single Quantum Coherence (HSQC) experiment. This work therefore aims to characterize the role of the LRR-LDLa linker in signal activation by assigning the residues within the linker and within relaxin that are involved in the interaction. Elucidation of the molecular mechanism of RXFP1 receptor activation including the involvement of the linker is essential for designing small molecules as RXFP receptor modulators, for effective treatment of cardiovascular diseases.