Molecular mechanism of Gαi activation by non-GPCR proteins with a Gα-Binding and Activating motif

• Published in: Nature communications 2017-05, Vol.8 (1), p.15163-15163, Article 15163
• Main Authors: de Opakua, Alain Ibáñez, Parag-Sharma, Kshitij, DiGiacomo, Vincent, Merino, Nekane, Leyme, Anthony, Marivin, Arthur, Villate, Maider, Nguyen, Lien T., de la Cruz-Morcillo, Miguel Angel, Blanco-Canosa, Juan B., Ramachandran, Sekar, Baillie, George S., Cerione, Richard A., Blanco, Francisco J., Garcia-Marcos, Mikel
• Format: Article
• Language: English
• Subjects: 101/6; 140/131; 631/45/535/878/1263; 631/80/86/2363; 82/80; 82/83; 96/95; Amino Acid Motifs - physiology; Cell Line; Enzyme Activation - physiology; GTP-Binding Protein alpha Subunits, Gi-Go - metabolism; Guanosine Diphosphate - metabolism; HEK293 Cells; Humanities and Social Sciences; Humans; Microfilament Proteins - metabolism; multidisciplinary; Nuclear Magnetic Resonance, Biomolecular; Protein Binding - physiology; Receptors, G-Protein-Coupled - metabolism; Science; Science (multidisciplinary); Signal Transduction - physiology; Vesicular Transport Proteins - metabolism
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/ncomms15163

Heterotrimeric G proteins are quintessential signalling switches activated by nucleotide exchange on Gα. Although activation is predominantly carried out by G-protein-coupled receptors (GPCRs), non-receptor guanine-nucleotide exchange factors (GEFs) have emerged as critical signalling molecules and therapeutic targets. Here we characterize the molecular mechanism of G-protein activation by a family of non-receptor GEFs containing a Gα-binding and -activating (GBA) motif. We combine NMR spectroscopy, computational modelling and biochemistry to map changes in Gα caused by binding of GBA proteins with residue-level resolution. We find that the GBA motif binds to the SwitchII/α3 cleft of Gα and induces changes in the G-1/P-loop and G-2 boxes (involved in phosphate binding), but not in the G-4/G-5 boxes (guanine binding). Our findings reveal that G-protein-binding and activation mechanisms are fundamentally different between GBA proteins and GPCRs, and that GEF-mediated perturbation of nucleotide phosphate binding is sufficient for Gα activation. Nonreceptor guanine-nucleotide exchange factors (GEFs) are emerging as important regulators of heterotrimeric G proteins. Here, the authors present structural and mechanistic insights into how a class of nonreceptor GEFs containing the Ga-Binding and Activating motif interact and modulate G proteins.