Met174 side chain is the site of photoinsertion of a substance P competitive peptide antagonist photoreactive in position 8.
Numerous photoaffinity studies of the NK-1 receptor have been carried out with peptide agonist analogues of substance P (SP). However, no information is available with regard to the domain interaction of peptide antagonists within this receptor. We describe herein the photoaffinity labelling of the SP receptor with a peptide antagonist analogue, Bapa(0)[(pBzl)Phe(8),DPro(9),MePhe(10),Trp(CHO)(11)]SP. Photolabelling, enzymatic or chemical cleavage of the covalent complex, purification via streptavidin-coated beads and matrix-assisted laser desorption/ionization time of flight mass spectrometry analysis led us to show that the methyl of Met174 side chain, within the receptor's second extracellular loop, is covalently linked to the antagonist photoreactive at position 8.
Sachon
E
E
UMR 7613 Université Pierre et Marie Curie, CNRS, case courrier 182, 4 place Jussieu, 75005 Paris Cedex, France.
Bolbach
G
G
Lavielle
S
S
Karoyan
P
P
Sagan
S
S
D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D006224:Cricetinae; D003488:Cyanogen Bromide / Q000494:pharmacology; D008024:Ligands; D008027:Light; D008715:Methionine / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D018040:Receptors, Neurokinin-1 / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013373:Substance P / Q000737:chemistry*
D008024:Ligands; D010455:Peptides; D018040:Receptors, Neurokinin-1; D013373:Substance P; D008715:Methionine; D003488:Cyanogen Bromide