Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2020 Oct 22;11(11):2336-2340.
doi: 10.1021/acsmedchemlett.0c00456. eCollection 2020 Nov 12.

Effects of C-Terminal B-Chain Modifications in a Relaxin 3 Agonist Analogue

Praveen Praveen  1 , Julien Tailhades  2   3 , K Johan Rosengren  4 , Mengjie Liu  1 , John D Wade  1   5 , Ross A D Bathgate  1   5 , Mohammed Akhter Hossain  1   5
Affiliations

Effects of C-Terminal B-Chain Modifications in a Relaxin 3 Agonist Analogue

Praveen Praveen et al. ACS Med Chem Lett. .

Abstract

The receptor for the neuropeptide relaxin 3, relaxin family peptide 3 (RXFP3) receptor, is an attractive pharmacological target for the control of eating, addictive, and psychiatric behaviors. Several structure-activity relationship studies on both human relaxin 3 (containing 3 disulfide bonds) and its analogue A2 (two disulfide bonds) suggest that the C-terminal carboxylic acid of the tryptophan residue in the B-chain is important for RXFP3 activity. In this study, we have added amide, alcohol, carbamate, and ester functionalities to the C-terminus of A2 and compared their structures and functions. As expected, the C-terminal amide form of A2 showed lower binding affinity for RXFP3 while ester and alcohol substitutions also demonstrated lower affinity. However, while these analogues showed slightly lower binding affinity, there was no significant difference in activation of RXFP3 compared to A2 bearing a C-terminal carboxylic acid, suggesting the binding pocket is able to accommodate additional atoms.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing financial interest.

Figures

Figure 1
Figure 1
Human relaxin-3 representation based on the NMR data highlighting all the key residues. Purple, A-chain; blue, B-chain highlighting the amino acids involved in RXFP3 binding; red, the C-terminal dipeptide important in RXFP3 activation; yellow, disulfide bridges.
Scheme 1
Scheme 1. Solid Phase Peptide Synthesis and Regioselective Disulfide Bridge Formation to Generate A2 Analogues (Yields 3-15%; Table S1)
Synthetic conditions: (i) 20% piperidine in DMF, 20 min, RT; (ii) Fmoc-AA-OH, HCTU, DIEA, DMF, 1 h, RT; (iii) TFA/DODT/TIS/H2O with 10 equiv of DSPS, 2 h, RT; (iii)′ TFA/DODT/TIS/H2O, 2h, RT; (iv) PBS/ACN (80:20) at pH 7.4, 2 h, RT; (v) addition of A-chain in 6 M GnHCl (pH 8.5) to B-chain in 6 M GnHCl (pH 5.5), 15 min, RT; (vi) I2, AcOH, 1 h, RT.
Figure 2
Figure 2
Competition binding (A) and cAMP activity assays of A2 and its analogues in (B) CHO-RXFP3 cells and (C) CHO-RXFP4 cells. Europium-labeled H3 B1-22R was used as the labeled ligand in the competition binding assays.
Figure 3
Figure 3
Circular dichroism spectroscopy overlapping showing that the overall secondary structure is maintained despite the C-terminal B-chain modification.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Patil N. A.; Rosengren K. J.; Separovic F.; Wade J. D.; Bathgate R. A. D.; Hossain M. A. Relaxin family peptides: structure–activity relationship studies. Br. J. Pharmacol. 2017, 174 (10), 950–961. 10.1111/bph.13684. - DOI - PMC - PubMed
    1. Praveen P.; Kocan M.; Valkovic A.; Bathgate R.; Hossain M. A. Single chain peptide agonists of relaxin receptors. Mol. Cell. Endocrinol. 2019, 487, 34–39. 10.1016/j.mce.2019.01.008. - DOI - PubMed
    1. Bathgate R. A. D.; van der Westhuizen M. L.; Halls E. T.; Callander G. E.; Kocan M.; Summers R. J. Relaxin family peptides and their receptors. Physiol. Rev. 2013, 93 (1), 405.10.1152/physrev.00001.2012. - DOI - PubMed
    1. Bathgate R. A. D.; Samuel C. S.; Burazin T. C. D.; Layfield S.; Claasz A. A.; Reytomas I. G. T.; Dawson N. F.; Zhao C.; Bond C.; Summers R. J.; Parry L. J.; Wade J. D.; Tregear G. W. Human Relaxin Gene 3 (H3) and the Equivalent Mouse Relaxin (M3) Gene: Novel members of the relaxin peptide family. J. Biol. Chem. 2002, 277 (2), 1148–1157. 10.1074/jbc.M107882200. - DOI - PubMed
    1. Smith C.; Walker A.; Hosken I.; Chua B.; Zhang C.; Haidar M.; Gundlach A. Relaxin-3/RXFP3 networks: an emerging target for the treatment of depression and other neuropsychiatric diseases?. Front. Pharmacol. 2014, 5, 46.10.3389/fphar.2014.00046. - DOI - PMC - PubMed