Premium
The electrostatic interactions of relaxin‐3 with receptor RXFP 4 and the influence of its B ‐chain C ‐terminal conformation
Author(s) -
Wang XinYi,
Guo YuQi,
Zhang WeiJie,
Shao XiaoXia,
Liu YaLi,
Xu ZengGuang,
Guo ZhanYun
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12830
Subject(s) - relaxin , receptor , mutant , peptide , chemistry , c terminus , mutagenesis , n terminus , biochemistry , microbiology and biotechnology , peptide sequence , biophysics , stereochemistry , biology , amino acid , gene
Relaxin‐3 (also known as insulin‐like peptide 7) is an insulin/relaxin‐superfamily peptide hormone that can bind and activate three relaxin‐family peptide receptors: RXFP 3, RXFP 4, and RXFP 1. Recently, we identified key electrostatic interactions between relaxin‐3 and its cognate receptor RXFP 3 by using a charge‐exchange mutagenesis approach. In the present study, the electrostatic interactions between relaxin‐3 and RXFP 4 were investigated with the same approach. Mutagenesis of the negatively charged extracellular residues of human RXFP 4 identified a conserved EXXXD (100–104) motif that is essential for RXFP 4 activation by relaxin‐3. Mutagenesis of the conserved positively charged Arg residues of relaxin‐3 demonstrated that B12Arg, B16Arg and B26Arg were all involved in the binding and activation of RXFP 4, especially B26Arg. The activity complementation between the mutant ligands and the mutant receptors suggested two probable electrostatic interaction pairs: Glu100 of RXFP 4 versus B26Arg of relaxin‐3, and Asp104 of RXFP 4 versus both B12Arg and B16Arg of relaxin‐3. For interaction with the essential EXXXD motifs of both RXFP 3 and RXFP 4, a folding‐back conformation of the relaxin‐3 B‐chain C‐terminus seems to be critical, because it brings B26Arg sufficiently close to B12Arg and B16Arg. To test this hypothesis, we replaced the conserved B23Gly‐B24Gly dipeptide of relaxin‐3 with an Ala‐Ser dipeptide that occupied the corresponding position of insulin‐like peptide 5 and resulted in an extended helical conformation. The mutant relaxin‐3 showed a significant decrease in receptor‐activation potency towards both RXFP 3 and RXFP 4, suggesting that a folding‐back conformation of the B‐chain C‐terminus was important for relaxin‐3 to efficiently interact with the EXXXD motifs of both receptors. Structured digital abstractRXFP4 and relaxin-3 physically interact by competition binding ( 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 )