Evidence for a direct interaction between the Thr11 residue of vasoactive intestinal polypeptide and Tyr184 located in the first extracellular loop of the VPAC2 receptor.
We developed previously VPAC(1) [vasoactive intestinal peptide (VIP)/pituitary adenylate cyclase-activating peptide (PACAP) receptor]>VPAC(2) receptor selective ligands. Replacement of the VIP-Thr(11) by an Arg(11) in these ligands contributed to their selectivity: Arg(11)-VIP had a 200-fold lower affinity when compared with VIP at VPAC(2) receptors as opposed to 3- to 5-fold higher affinity at VPAC(1) receptors. Comparison of the binding and functional properties of related VIP analogues suggested that the VPAC(1) selectivity of Arg(11)-VIP was due to the loss of a hydrogen bond between the hydroxy group of Thr residue and the VPAC(2) receptor, steric hindrance between the Arg side chain and the VPAC(2) receptor and charge attraction by the VPAC(1) receptor. Comparison of the ability of VIP analogues to activate adenylate cyclase through chimaeric VPAC(1)/VPAC(2) and VPAC(2)/VPAC(1) receptors indicated that the first extracellular receptor loop carried most of the VPAC(2) receptors' ability to discriminate VIP from Arg(11)-VIP. Based on results obtained for a truncated VPAC(2) receptor and the closely related PACAP-preferring receptor (PAC(1)) and secretin receptors, we hypothesized that Thr(11) interacted with the VPAC(2) receptor Tyr(184) (similar to the VPAC(1) receptor Phe(200) residue). The Y184F (Tyr(184)-->Phe) VPAC(2) mutant lost the ability to discriminate VIP from Val(11)-VIP, and the F200Y VPAC(1) mutant acquired the ability to discriminate the natural peptide from Val(11)-VIP. These results support the hypothesis that the hydroxy group of the native VIP-Thr(11) side chain can indeed form a hydrogen bond with the Tyr side chain in the VPAC(2) receptor.
Nachtergael
Ingrid
I
Department of Biochemistry and Nutrition, School of Medicine, Université Libre de Bruxelles, Bât G/E, CP 611, 808 route de Lennik, B-1070 Bruxelles, Belgium.
Vertongen
Pascale
P
Langer
Ingrid
I
Perret
Jason
J
Robberecht
Patrick
P
Waelbroeck
Magali
M
D000262:Adenylyl Cyclases / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005765:Gastrointestinal Agents / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D018005:Receptors, Vasoactive Intestinal Peptide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051239:Receptors, Vasoactive Intestinal Peptide, Type II; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013912:Threonine / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D014660:Vasoactive Intestinal Peptide / Q000737:chemistry* / Q000378:metabolism*
D005765:Gastrointestinal Agents; D010446:Peptide Fragments; D018005:Receptors, Vasoactive Intestinal Peptide; D051239:Receptors, Vasoactive Intestinal Peptide, Type II; D011993:Recombinant Fusion Proteins; D013912:Threonine; D014660:Vasoactive Intestinal Peptide; D014443:Tyrosine; D000262:Adenylyl Cyclases