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The complex binding mode of the peptide hormone H2 relaxin to its receptor RXFP1.

Sethi A, Bruell S, Patil N, Hossain MA, Scott DJ, Petrie EJ, Bathgate RA, Gooley PR - Nat Commun (2016)

Bottom Line: H2 relaxin is hypothesized to bind with high affinity to the LRR domain enabling the LDLa module to bind and activate the transmembrane domain of RXFP1.Here we define a relaxin-binding site on the LDLa-LRR linker, essential for the high affinity of H2 relaxin for the ectodomain of RXFP1, and show that residues within the LDLa-LRR linker are critical for receptor activation.We propose H2 relaxin binds and stabilizes a helical conformation of the LDLa-LRR linker that positions residues of both the linker and the LDLa module to bind the transmembrane domain and activate RXFP1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry &Molecular Biology, The University of Melbourne, Victoria 3010, Australia.

ABSTRACT
H2 relaxin activates the relaxin family peptide receptor-1 (RXFP1), a class A G-protein coupled receptor, by a poorly understood mechanism. The ectodomain of RXFP1 comprises an N-terminal LDLa module, essential for activation, tethered to a leucine-rich repeat (LRR) domain by a 32-residue linker. H2 relaxin is hypothesized to bind with high affinity to the LRR domain enabling the LDLa module to bind and activate the transmembrane domain of RXFP1. Here we define a relaxin-binding site on the LDLa-LRR linker, essential for the high affinity of H2 relaxin for the ectodomain of RXFP1, and show that residues within the LDLa-LRR linker are critical for receptor activation. We propose H2 relaxin binds and stabilizes a helical conformation of the LDLa-LRR linker that positions residues of both the linker and the LDLa module to bind the transmembrane domain and activate RXFP1.

No MeSH data available.


Mechanism of H2 relaxin binding to RXFP1.(a) Cartoon model of the domain structure of RXFP1. (b) H2 relaxin interacts and binds to the linker and LRR domain of RXFP1. (c) Binding of H2 relaxin stabilizes and extends a helix within the linker to orient and enable interactions of the LDLa module and residues within the linker to exoloop-2 of the TMD to facilitate receptor activation. The TMD (blue), LRR domain (grey), LRR-LDLa linker (magenta) and LDLa module (cyan) of RXFP1 are indicated. Additional loops are coloured red with exoloop-2 (EL2) of the TMD annotated. H2 relaxin is coloured green.
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f8: Mechanism of H2 relaxin binding to RXFP1.(a) Cartoon model of the domain structure of RXFP1. (b) H2 relaxin interacts and binds to the linker and LRR domain of RXFP1. (c) Binding of H2 relaxin stabilizes and extends a helix within the linker to orient and enable interactions of the LDLa module and residues within the linker to exoloop-2 of the TMD to facilitate receptor activation. The TMD (blue), LRR domain (grey), LRR-LDLa linker (magenta) and LDLa module (cyan) of RXFP1 are indicated. Additional loops are coloured red with exoloop-2 (EL2) of the TMD annotated. H2 relaxin is coloured green.

Mentions: Consequently, we propose the functions of the residues of the linker as follows: residues within the sequence GDNNGW directly interact with EL2 of the TMD and together with the LDLa module1012 are important in inducing the activated state of the receptor. Mutation of these residues results in loss of interaction with EL2 and a modest loss of H2 relaxin binding. While Gly45 and Trp46 may directly contact H2 relaxin, mutation of these residues shows that they also have important roles in maintaining the structure of the linker to bind H2 relaxin. Their mutation therefore has a combined effect on both H2 relaxin binding and receptor activation. Asp42 does not appear to have such a role in H2 relaxin binding, as the mutant G41A/D42A-RXFP1(1–72) appears to bind H2 relaxin relatively normally, and the reasons for the significant loss of H2 relaxin affinity on mutation in RXFP1 are not clear. Phe50, Phe54 and Tyr58 are proposed to largely form the H2 relaxin-binding epitope. The mechanism we now propose is that H2 relaxin binds to the LRR via B-chain residues and to the linker via A-chain residues resulting in a high-affinity complex (Fig. 8). Binding of H2 relaxin simultaneously stabilizes and extends a helical conformational state of the linker. This conformational change positions residues of the linker, especially Gly41 to Trp46, and also residues of the N-terminal region of the LDLa module1012 and H2 relaxin to form an interaction with the TMD resulting in receptor activation and the appropriate signalling pathways1012142829.


The complex binding mode of the peptide hormone H2 relaxin to its receptor RXFP1.

Sethi A, Bruell S, Patil N, Hossain MA, Scott DJ, Petrie EJ, Bathgate RA, Gooley PR - Nat Commun (2016)

Mechanism of H2 relaxin binding to RXFP1.(a) Cartoon model of the domain structure of RXFP1. (b) H2 relaxin interacts and binds to the linker and LRR domain of RXFP1. (c) Binding of H2 relaxin stabilizes and extends a helix within the linker to orient and enable interactions of the LDLa module and residues within the linker to exoloop-2 of the TMD to facilitate receptor activation. The TMD (blue), LRR domain (grey), LRR-LDLa linker (magenta) and LDLa module (cyan) of RXFP1 are indicated. Additional loops are coloured red with exoloop-2 (EL2) of the TMD annotated. H2 relaxin is coloured green.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4837482&req=5

f8: Mechanism of H2 relaxin binding to RXFP1.(a) Cartoon model of the domain structure of RXFP1. (b) H2 relaxin interacts and binds to the linker and LRR domain of RXFP1. (c) Binding of H2 relaxin stabilizes and extends a helix within the linker to orient and enable interactions of the LDLa module and residues within the linker to exoloop-2 of the TMD to facilitate receptor activation. The TMD (blue), LRR domain (grey), LRR-LDLa linker (magenta) and LDLa module (cyan) of RXFP1 are indicated. Additional loops are coloured red with exoloop-2 (EL2) of the TMD annotated. H2 relaxin is coloured green.
Mentions: Consequently, we propose the functions of the residues of the linker as follows: residues within the sequence GDNNGW directly interact with EL2 of the TMD and together with the LDLa module1012 are important in inducing the activated state of the receptor. Mutation of these residues results in loss of interaction with EL2 and a modest loss of H2 relaxin binding. While Gly45 and Trp46 may directly contact H2 relaxin, mutation of these residues shows that they also have important roles in maintaining the structure of the linker to bind H2 relaxin. Their mutation therefore has a combined effect on both H2 relaxin binding and receptor activation. Asp42 does not appear to have such a role in H2 relaxin binding, as the mutant G41A/D42A-RXFP1(1–72) appears to bind H2 relaxin relatively normally, and the reasons for the significant loss of H2 relaxin affinity on mutation in RXFP1 are not clear. Phe50, Phe54 and Tyr58 are proposed to largely form the H2 relaxin-binding epitope. The mechanism we now propose is that H2 relaxin binds to the LRR via B-chain residues and to the linker via A-chain residues resulting in a high-affinity complex (Fig. 8). Binding of H2 relaxin simultaneously stabilizes and extends a helical conformational state of the linker. This conformational change positions residues of the linker, especially Gly41 to Trp46, and also residues of the N-terminal region of the LDLa module1012 and H2 relaxin to form an interaction with the TMD resulting in receptor activation and the appropriate signalling pathways1012142829.

Bottom Line: H2 relaxin is hypothesized to bind with high affinity to the LRR domain enabling the LDLa module to bind and activate the transmembrane domain of RXFP1.Here we define a relaxin-binding site on the LDLa-LRR linker, essential for the high affinity of H2 relaxin for the ectodomain of RXFP1, and show that residues within the LDLa-LRR linker are critical for receptor activation.We propose H2 relaxin binds and stabilizes a helical conformation of the LDLa-LRR linker that positions residues of both the linker and the LDLa module to bind the transmembrane domain and activate RXFP1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry &Molecular Biology, The University of Melbourne, Victoria 3010, Australia.

ABSTRACT
H2 relaxin activates the relaxin family peptide receptor-1 (RXFP1), a class A G-protein coupled receptor, by a poorly understood mechanism. The ectodomain of RXFP1 comprises an N-terminal LDLa module, essential for activation, tethered to a leucine-rich repeat (LRR) domain by a 32-residue linker. H2 relaxin is hypothesized to bind with high affinity to the LRR domain enabling the LDLa module to bind and activate the transmembrane domain of RXFP1. Here we define a relaxin-binding site on the LDLa-LRR linker, essential for the high affinity of H2 relaxin for the ectodomain of RXFP1, and show that residues within the LDLa-LRR linker are critical for receptor activation. We propose H2 relaxin binds and stabilizes a helical conformation of the LDLa-LRR linker that positions residues of both the linker and the LDLa module to bind the transmembrane domain and activate RXFP1.

No MeSH data available.