Large-scale conformational rearrangement of the {alpha}5-helix of G{alpha} subunits in complex with the guanine nucleotide exchange factor Ric8A [Protein Structure and Folding] (Journal of Biological Chemistry)

Resistance to inhibitors of cholinesterase 8A (Ric8A) protein is an important G protein-coupled receptor (GPCR)-independent regulator of G protein ?-subunits (G?), acting as a guanine nucleotide exchange factor (GEF) and a chaperone. Insights into the complex between Ric8A and G? hold the key to understanding the mechanisms underlying noncanonical activation of G-protein signaling as well as the folding of nascent G? proteins. Here, we examined the structure of the complex of Ric8A with minimized G?i (miniG?i) in solution by small-angle X-ray scattering (SAXS) and exploited the scattering profile in modeling of the Ric8A/miniG?i complex by steered molecular dynamics (SMD) simulations. A small set of models of the complex featured minimal clash scores, excellent agreement with the experimental SAXS data, and a large-scale rearrangement of the signal-transducing ?5-helix of G? away from its ?-sheet core. The resulting interface involved the G? ?5-helix bound to the concave surface of Ric8A and the G? ?-sheet that wraps around the C-terminal part of the Ric8A armadillo domain, leading to a severe disruption of the GDP-binding site. Further modeling of the flexible C-terminal tail of Ric8A indicated that it interacts with the effector surface of G?. This smaller interface may enable the Ric8A-bound G? to interact with GTP. The two-interface interaction with G? described here distinguishes Ric8A from GPCRs and non-GPCR regulators of G-protein signaling.