Molecular properties of endogenous RFamide-related peptide-3 and its interaction with receptors.
Based on database searches of DNA sequences, we previously reported a gene encoding peptides possessing Arg-Phe-NH(2) (RFamide) at their C termini. This gene, RFamide-related peptide (RFRP), was expected to encode several different peptides (i.e., RFRP-1, -2, and -3). In the present study, we purified endogenous RFRP-3 from bovine hypothalamus, and demonstrated that it consisted of 28 amino acid residues. After constructing a sandwich enzyme immunoassay for RFRP-3, we analyzed the tissue distribution of endogenous RFRP-3 in rats and found its concentration to be highest in the hypothalamus. In binding assays, [125I]-labeled RFRP-3 bound to OT7T022 with high affinity, but its binding affinity to HLWAR77 was low. On the other hand, [125I]-labeled neuropeptide FF (NPFF) bound to both OT7T022 and HLWAR77 with high affinity. By serial deletion in the N-terminal portions of RFRP-3 and NPFF, we found that four C-terminal amino acid residues (i.e., PQRFamide), which were common between the two peptides, comprised a core sequence responsible for binding with the receptors, whereas three amino acid residues (i.e., PNL in RFRP-3 and LFQ in NPFF) added to the N terminus of PQRFamide played crucial roles in the agonistic activities of RFRP-3 and NPFF for OT7T022 and HLWAR77, respectively.
Yoshida
Hiromi
H
Discovery Research Laboratories I, Pharmaceutical Research Division, Takeda Chemical Industries, Ltd., 10 Wadai, Ibaraki 300-4293, Tsukuba, Japan.
Habata
Yugo
Y
Hosoya
Masaki
M
Kawamata
Yuji
Y
Kitada
Chieko
C
Hinuma
Shuji
S
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D001665:Binding Sites; D001667:Binding, Competitive; D002417:Cattle; D002845:Chromatography; D006801:Humans; D007031:Hypothalamus / Q000737:chemistry / Q000378:metabolism; D007124:Immunoenzyme Techniques; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009479:Neuropeptides / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D009842:Oligopeptides / Q000378:metabolism; D051381:Rats; D018013:Receptors, Neuropeptide / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization; D013788:Thalamus / Q000737:chemistry / Q000378:metabolism
D000906:Antibodies; D009479:Neuropeptides; D009842:Oligopeptides; C416987:RFamide peptide; D018013:Receptors, Neuropeptide; C083929:neuropeptide FF receptor; C053651:phenylalanyl-leucyl-phenylalanyl-glutaminyl-prolyl-glutaminyl-arginyl-phenylalaninamide