The common C-terminal sequences of substance P and neurokinin A contact the same region of the NK-1 receptor.
Although neurokinin A (NKA), a tachykinin peptide with sequence homology to substance P (SP), is a weak competitor of radiolabeled SP binding to the NK-1 receptor (NK-1R), more recent direct binding studies using radiolabeled NKA have demonstrated an unexpected high-affinity interaction with this receptor. To document the site of interaction between NKA and the NK-1R, we have used a photoreactive analogue of NKA containing p-benzoyl-L-phenylalanine (Bpa) substituted in position 7 of the peptide. Peptide mapping studies of the receptor photolabeled by (125)I-iodohistidyl(1)-Bpa(7)NKA have established that the site of photoinsertion is located within a segment of the receptor extending from residues 178 to 190 (VVCMIEWPEHPNR). We have previously shown that (125)I-BH-Bpa(8)SP, a photoreactive analogue of SP, covalently attaches to M(181) within this same receptor sequence. Importantly, both of these peptides ((125)I-iodohistidyl(1)-Bpa(7)NKA and (125)I-BH-Bpa(8)SP) have the photoreactive amino acid in an equivalent position within the conserved tachykinin carboxyl-terminal tail. In this report, we also show that site-directed mutagenesis of M(181) to A(181) in the NK-1R results in a complete loss of photolabeling of both peptides to this receptor site, indicating that the equivalent position of SP and NKA, when bound to the NK-1R, contact the same residue.
Bremer
A A
AA
Department of Pharmacology and Experimental Therapeutics, Boston University School of Medicine, L-611, 80 E. Concord Street, Boston, MA 02118, USA.
Leeman
S E
SE
Boyd
N D
ND
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D017124:Conserved Sequence; D006224:Cricetinae; D020128:Inhibitory Concentration 50; D008024:Ligands; D009154:Mutation / Q000235:genetics; D015288:Neurokinin A / Q000737:chemistry* / Q000378:metabolism*; D010449:Peptide Mapping; D019920:Photoaffinity Labels; D011485:Protein Binding; D011487:Protein Conformation; D018040:Receptors, Neurokinin-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013373:Substance P / Q000737:chemistry* / Q000378:metabolism*; D014162:Transfection; D014357:Trypsin / Q000378:metabolism
D008024:Ligands; D019920:Photoaffinity Labels; D018040:Receptors, Neurokinin-1; D013373:Substance P; D015288:Neurokinin A; D014357:Trypsin