A novel substance P binding site in bovine adrenal medulla.
Radioligand binding techniques were used to characterize the substance P (SP) binding site on membranes prepared from bovine adrenal medullae. 125I-labelled Bolton-Hunter substance P (BHSP), which recognises the C-terminally directed, SP-preferring NK1 receptor, showed no specific binding. In contrast, binding of [3H]SP was saturable (at 6 nM) and reversible, with an equilibrium dissociation constant (Kd) 1.46 +/- 0.73 nM, Bmax 0.73 +/- 0.06 pmol/g wet weight and Hill coefficient 0.98 +/- 0.01. Specific binding of [3H]SP was displaced by SP greater than neurokinin A (NKA) greater than SP(3-11) approximately SP(1-9) greater than SP(1-7) approximately SP(1-4) approximately SP(1-6), with neurokinin B (NKB) and SP(1-3) very weak competitors and SP(5-11), SP(7-11) and SP(9-11) causing negligible inhibition (up to 10 microM). This potency order is quite distinct from that seen with binding to an NK1 site, a conclusion confirmed by the lack of BHSP binding. It appears that Lys3 and/or Pro4 are critical for binding, suggesting an anionic binding site. These data suggest the existence of an unusual binding site which may represent a novel SP receptor. This site appears to require the entire sequence of the SP molecule for full recognition.
Geraghty
D P
DP
Department of Biological Sciences, Deakin University, Vic., Australia.
Livett
B G
BG
Rogerson
F M
FM
Burcher
E
E
D000313:Adrenal Medulla / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002417:Cattle; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism*; D018040:Receptors, Neurokinin-1; D017981:Receptors, Neurotransmitter / Q000187:drug effects / Q000378:metabolism*; D013373:Substance P / Q000378:metabolism*
D010446:Peptide Fragments; D018040:Receptors, Neurokinin-1; D017981:Receptors, Neurotransmitter; D013373:Substance P