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Mutational analysis of the human vasoactive intestinal peptide receptor subtype VPAC 2 : role of basic residues in the second transmembrane helix
Author(s) -
Vertongen P,
Solano R M,
Perret J,
Langer I,
Robberecht P,
Waelbroeck M
Publication year - 2001
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1038/sj.bjp.0704195
Subject(s) - vasoactive intestinal peptide , receptor , transmembrane domain , ligand (biochemistry) , biology , adenylate kinase , chemistry , endocrinology , biochemistry , medicine , neuropeptide
We investigated the role of two conserved basic residues in the second transmembrane helix arginine 172 (R172) and lysine 179 (K179) of the VPAC 2 receptor. Vasoactive intestinal polypeptide (VIP) activated VPAC 2 receptors with an EC 50 value of 7 n M , as compared to 150, 190 and 4000 n M at R172L, R172Q and K179Q‐VPAC 2 receptors, respectively. It was inactive at K179I mutated VPAC 2 receptors. These results suggested that both basic residues were probably implicated in receptor recognition and activation. The VPAC 2 ‐selective VIP analogue, [hexanoyl‐His 1 ]‐VIP (C 6 ‐VIP), had a higher affinity and efficacy as compared to VIP at the mutated receptors. VIP, Asn 3 ‐VIP and Gln 3 ‐VIP activated adenylate cyclase through R172Q receptors with EC 50 values of 190, 2 and 2 n M , respectively, and through R172L receptors with EC 50 values of 150, 12 and 8 n M , respectively. Asn 3 ‐VIP and Gln 3 ‐VIP behaved as partial agonists at the wild type receptor, with E max values (in per cent of VIP) of 75 and 52%, respectively. In contrast, they were more efficient than VIP (E max values of 150 and 150% at the R172Q VPAC 2 receptors, and of 400 and 360% at the R172L receptors, respectively). These results suggested that the receptor's R172 and the ligand's aspartate 3 are brought in close proximity in the active ligand‐receptor complex. The K179I and K179Q mutated receptors had a lower affinity than the wild‐type receptors for all the agonists tested in this work: we were unable to identify the VIP amino acid(s) that interact with K179.British Journal of Pharmacology (2001) 133 , 1249–1254; doi: 10.1038/sj.bjp.0704195