Abstract
The VPAC1 receptor belongs to family B of G protein-coupled receptors (GPCR-B) and is activated upon binding of the vasoactive intestinal peptide (VIP). Despite the recent determination of the structure of the N terminus of several members of this receptor family, little is known about the structure of the transmembrane (TM) region and about the molecular mechanisms leading to activation. In the present study, we designed a new structural model of the TM domain and combined it with experimental mutagenesis experiments to investigate the interaction network that governs ligand binding and receptor activation. Our results suggest that this network involves the cluster of residues Arg188 in TM2, Gln380 in TM7, and Asn229 in TM3. This cluster is expected to be altered upon VIP binding, because Arg188 has been shown previously to interact with Asp3 of VIP. Several point mutations at positions 188, 229, and 380 were experimentally characterized and were shown to severely affect VIP binding and/or VIP-mediated cAMP production. Double mutants built from reciprocal residue exchanges exhibit strong cooperative or anticooperative effects, thereby indicating the spatial proximity of residues Arg188, Gln380, and Asn229. Because these residues are highly conserved in the GPCR-B family, they can moreover be expected to have a general role in mediating function.
Footnotes
↵ The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
This work was supported by the Brussels Region TheraVip project; the Belgian State Science Policy Office through an Interuniversity Attraction Poles Programme (DYSCO); the Belgian Fund for Scientific Research [Grant 3.4553.06]; and the President of Russian Federation [Grant MK-125.2008.4].
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
doi:10.1124/mol.110.063578.
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ABBREVIATIONS:
- GPCR
- G protein-coupled receptor
- 3D
- three dimensional
- Aln
- alignment
- MD
- molecular dynamics
- OPSD
- bovine visual rhodopsin
- TM
- transmembrane
- VIP
- vasoactive intestinal peptide
- wt
- wild type
- PTH
- parathyroid hormone.
- Received January 13, 2010.
- Accepted June 23, 2010.
- Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics
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