Abstract
Vasoactive intestinal polypeptide receptor (VIP1R) is a widely expressed class B G protein-coupled receptor and a drug target for the treatment of neuronal, metabolic, and inflammatory diseases. However, our understanding of its mechanism of action and the potential of drug discovery targeting this receptor is limited by the lack of structural information of VIP1R. Here we report a cryo-electron microscopy structure of human VIP1R bound to PACAP27 and Gs heterotrimer, whose complex assembly is stabilized by a NanoBiT tethering strategy. Comparison with other class B GPCR structures reveals that PACAP27 engages VIP1R with its N-terminus inserting into the ligand binding pocket at the transmembrane bundle of the receptor, which subsequently couples to the G protein in a receptor-specific manner. This structure has provided insights into the molecular basis of PACAP27 binding and VIP receptor activation. The methodology of the NanoBiT tethering may help to provide structural information of unstable complexes.
Highlights
Vasoactive intestinal polypeptide receptors, known as vasoactive intestinal peptide (VIP) receptors, including VIP1R and VIP2R, belong to the class B1 of G protein-coupled receptors
We demonstrate that the NanoBiT tethering method can be applied to other GPCR–G protein complexes
When the NanoBiT is dissected between residues 156 and 157, it can be split into a large component containing 156 amino acid residues named large BiT (LgBiT), and a 13-amino acid peptide called small BiT (SmBiT, Supplementary Fig. 1a)
Summary
Vasoactive intestinal polypeptide receptors, known as VIP receptors, including VIP1R and VIP2R, belong to the class B1 of G protein-coupled receptors. Various methods have been developed to improve the stability of GPCR-signal transducer complexes, such as the use of thermo-stabilizing mutations[15], nanobodies, and antibody fragments[16,17], to facilitate structural studies. We have developed a method to stabilize the interaction between VIP1R and the Gs heterotrimer by bringing the two proteins into close proximity through a NanoBiT tethering approach. This method greatly improved the stability and homogeneity of the PACAP27–VIP1R–Gs protein complex, allowing structural determination of human VIP1R in complex with PACAP27 and Gs heterotrimer. We demonstrate that the NanoBiT tethering method can be applied to other GPCR–G protein complexes
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