The structural and functional role of the B-chain C-terminal arginine in the relaxin-3 peptide antagonist, R3(BDelta23-27)R/I5.
Relaxin-3, a member of the insulin superfamily, is involved in regulating stress and feeding behavior. It is highly expressed in the brain and is the endogenous ligand for the receptor RXFP3. As relaxin-3 also interacts with the relaxin receptor RXFP1, selective agonists and antagonists are crucial for studying the physiological function(s) of the relaxin-3/RXFP3 pair. The analog R3(BDelta23-27)R/I5, in which a C-terminally truncated human relaxin-3 (H3) B-chain is combined with the INSL5 A-chain, is a potent selective RXFP3 antagonist and has an Arg residue remaining on the B-chain C-terminus as a consequence of the recombinant protein production process. To investigate the role of this residue in the RXFP3 receptor binding and activation, the analogs R3(BDelta23-27)R/I5 and R3(BDelta23-27)R containing the B-chain C-terminal Arg as well as R3(BDelta23-27)/I5 and R3(BDelta23-27), both lacking the Arg, were chemically assembled and their secondary structure and receptor activity assessed. The peptides generally had a similar conformation but those with the extra Arg residue displayed a significantly increased affinity for the RXFP3. Interestingly, in contrast to R3(BDelta23-27)R and R3(BDelta23-27)R/I5, the peptide R3(BDelta23-27) is a weak agonist. This suggests that the C-terminal Arg, although increasing the affinity, alters the manner in which the peptide binds to the receptor and thereby prevents activation, giving R3(BDelta23-27)R/I5 its potent antagonistic activity.
Hossain
Mohammed Akhter
MA
Howard Florey Institute, University of Melbourne, Melbourne, Victoria 3010, Australia.
Bathgate
Ross A D
RA
Rosengren
K Johan
KJ
Shabanpoor
Fazel
F
Zhang
Suode
S
Lin
Feng
F
Tregear
Geoffrey W
GW
Wade
John D
JD
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D006801:Humans; D007328:Insulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012065:Relaxin / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
D004220:Disulfides; D007328:Insulin; C084797:Leydig insulin-like protein; D010455:Peptides; D011506:Proteins; C480238:RLN3 protein, human; C410092:RXFP3 protein, human; D043562:Receptors, G-Protein-Coupled; D011993:Recombinant Fusion Proteins; D012065:Relaxin; D001120:Arginine; D000242:Cyclic AMP; D003545:Cysteine