1. Structure, function, and dynamics of the G alpha binding domain of Ric-8A
- Author
-
Makaía M. Papasergi-Scott, Gregory G. Tall, Baisen Zeng, Franz Hagn, Wenxi Yu, Tzanko Doukov, Tung-Chung Mou, Stephen R. Sprang, and Andrea Steiner
- Subjects
Models, Molecular ,Protein Conformation, alpha-Helical ,G protein ,Stereochemistry ,Recombinant Fusion Proteins ,Genetic Vectors ,Gene Expression ,GTP-Binding Protein alpha Subunits, Gi-Go ,Crystallography, X-Ray ,Article ,03 medical and health sciences ,Protein structure ,Structural Biology ,Heterotrimeric G protein ,Escherichia coli ,Animals ,Guanine Nucleotide Exchange Factors ,Histidine ,Protein Interaction Domains and Motifs ,Nucleotide ,Amino Acid Sequence ,Cloning, Molecular ,Molecular Biology ,Peptide sequence ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,Binding Sites ,Chemistry ,C-terminus ,030302 biochemistry & molecular biology ,Nuclear Proteins ,Rats ,Guanine Nucleotide Exchange Factor ,Heteronuclear Nuclear Magnetic Resonance ,Heterotrimeric G Protein ,Molecular Chaperone ,Protein Dynamics ,Protein Structure ,Small-angle X-ray Scattering ,X-ray Crystallography ,Mutation ,Protein Conformation, beta-Strand ,Guanosine Triphosphate ,Guanine nucleotide exchange factor ,Protein Multimerization ,Oligopeptides ,Protein Binding ,Binding domain - Abstract
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 (G alpha). We report the 2.2-angstrom crystal structure of the Ric-8A G alpha-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 G alpha-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 H-1-N-1(5)-TROSY spectra of [H-2, N-15]-labeled Gail in the presence of R452 reveals chemical shift perturbations of the C terminus and Gail residues involved in nucleotide binding.
- Published
- 2019