%0 Journal Article
%T A neurodevelopmental disorder mutation locks G proteins in the transitory pre-activated state.
%A Knight KM
%A Krumm BE
%A Kapolka NJ
%A Ludlam WG
%A Cui M
%A Mani S
%A Prytkova I
%A Obarow EG
%A Lefevre TJ
%A Wei W
%A Ma N
%A Huang XP
%A Fay JF
%A Vaidehi N
%A Smrcka AV
%A Slesinger PA
%A Logothetis DE
%A Martemyanov KA
%A Roth BL
%A Dohlman HG
%J Nat Commun
%V 15
%N 1
%D 2024 Aug 5
%M 39103320
%F 17.694
%R 10.1038/s41467-024-50964-z
%X Many neurotransmitter receptors activate G proteins through exchange of GDP for GTP. The intermediate nucleotide-free state has eluded characterization, due largely to its inherent instability. Here we characterize a G protein variant associated with a rare neurological disorder in humans. GαoK46E has a charge reversal that clashes with the phosphate groups of GDP and GTP. As anticipated, the purified protein binds poorly to guanine nucleotides yet retains wild-type affinity for G protein βγ subunits. In cells with physiological concentrations of nucleotide, GαoK46E forms a stable complex with receptors and Gβγ, impeding effector activation. Further, we demonstrate that the mutant can be easily purified in complex with dopamine-bound D2 receptors, and use cryo-electron microscopy to determine the structure, including both domains of Gαo, without nucleotide or stabilizing nanobodies. These findings reveal the molecular basis for the first committed step of G protein activation, establish a mechanistic basis for a neurological disorder, provide a simplified strategy to determine receptor-G protein structures, and a method to detect high affinity agonist binding in cells.