Structure, Function, and Dynamics of the Gα Binding Domain of Ric-8A.

Ric-8A is a 530-amino acid cytoplasmic molecular chaperone and guanine nucleotide exchange factor (GEF) for i, q, and 12/13 classes of heterortrimeric G protein alpha subunits (Gα). We report the 2.2-Å crystal structure of the Ric-8A Gα-binding domain with GEF activity, residues 1–452, and is phosphorylated at Ser435 and Thr440. Residues 1–429 adopt a superhelical fold comprised of Armadillo (ARM) and HEAT repeats, and the C terminus is disordered. One of the phosphorylated residues potentially binds to a basic cluster in an ARM motif. Amino acid sequence conservation and published hydrogen-deuterium exchange data indicate repeats 3 through 6 to be a putative Gα-binding surface. Normal mode modeling of small-angle X-ray scattering data indicates that phosphorylation induces relative rotation between repeats 1–4, 5–6, and 7–9. 2D 1H-15N-TROSY spectra of [2H,15N]-labeled Gαi1 in the presence of R452 reveals chemical shift perturbations of the C terminus and Gαi1 residues involved in nucleotide binding. • The G protein α subunit binding domain of Ric-8A is an Armadillo/HEAT repeat fold • Hydrogen-deuterium exchange data suggest a Gα binding surface on Ric-8A • Ric-8A binds elements of Gα also recognized by G protein-coupled receptors • The Ric-8A fold harbors a putative binding site for its C-terminal phosphoserine Zeng et al. use X-ray crystallography and small-angle scattering, in conjunction with NMR spectroscopy, to reveal the structure of and dynamics of the G protein chaperone and activator Ric-8A and probe its interaction with the G protein alpha subunit i1. [ABSTRACT FROM AUTHOR]