title: Probing Gαi1 protein activation at single-amino acid resolution.
abstract: We present comprehensive maps at single-amino acid resolution of the residues stabilizing the human Gαi1 subunit in nucleotide- and receptor-bound states. We generated these maps by measuring the effects of alanine mutations on the stability of Gαi1 and the rhodopsin-Gαi1 complex. We identified stabilization clusters in the GTPase and helical domains responsible for structural integrity and the conformational changes associated with activation. In activation cluster I, helices α1 and α5 pack against strands β1-β3 to stabilize the nucleotide-bound states. In the receptor-bound state, these interactions are replaced by interactions between α5 and strands β4-β6. Key residues in this cluster are Y320, which is crucial for the stabilization of the receptor-bound state, and F336, which stabilizes nucleotide-bound states. Destabilization of helix α1, caused by rearrangement of this activation cluster, leads to the weakening of the interdomain interface and release of GDP.
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.|Department of Biology, Swiss Federal Institute of Technology Zurich, Zurich, Switzerland.
authors.forename: Dawei
authors.initials: D
authors.lastname: Sun
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom.
authors.forename: Tilman
authors.initials: T
authors.lastname: Flock
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.|Condensed Matter Theory Group, Paul Scherrer Institut, Villigen, Switzerland.
authors.forename: Xavier
authors.initials: X
authors.lastname: Deupi
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.
authors.forename: Shoji
authors.initials: S
authors.lastname: Maeda
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.|Department of Biology, Swiss Federal Institute of Technology Zurich, Zurich, Switzerland.
authors.forename: Milos
authors.initials: M
authors.lastname: Matkovic
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.
authors.forename: Sandro
authors.initials: S
authors.lastname: Mendieta
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.|Department of Biology, Swiss Federal Institute of Technology Zurich, Zurich, Switzerland.
authors.forename: Daniel
authors.initials: D
authors.lastname: Mayer
authors.affiliation: F. Hoffmann-La Roche AG, Pharma Research & Early Development, Discovery Technologies, Basel, Switzerland.
authors.forename: Roger
authors.initials: R
authors.lastname: Dawson
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.|Department of Biology, Swiss Federal Institute of Technology Zurich, Zurich, Switzerland.
authors.forename: Gebhard F X
authors.initials: GFX
authors.lastname: Schertler
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom.
authors.forename: M
authors.initials: M
authors.lastname: Madan Babu
authors.affiliation: Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland.|Department of Biology, Swiss Federal Institute of Technology Zurich, Zurich, Switzerland.
authors.forename: Dmitry B
authors.initials: DB
authors.lastname: Veprintsev
mesh_terms: D000596:Amino Acids / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D004252:DNA Mutational Analysis; D044385:GTP-Binding Protein alpha Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D055550:Protein Stability; D012243:Rhodopsin / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D044385:GTP-Binding Protein alpha Subunits; D004247:DNA; D012243:Rhodopsin