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- PDB-8el8: CryoEM structure of Resistance to Inhibitors of Cholinesterase-8B... -

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Basic information

Entry
Database: PDB / ID: 8el8
TitleCryoEM structure of Resistance to Inhibitors of Cholinesterase-8B (Ric-8B) in complex with olfactory G protein alpha olf
Components
  • Guanine nucleotide-binding protein G(olf) subunit alpha
  • Isoform 1 of Synembryn-B
KeywordsSIGNALING PROTEIN / chaperone / armadillo repeat / GEF / Ras
Function / homology
Function and homology information


Adenylate cyclase activating pathway / sensory perception of chemical stimulus / response to caffeine / G-protein alpha-subunit binding / Adenylate cyclase inhibitory pathway / adenylate cyclase regulator activity / protein folding chaperone / guanyl-nucleotide exchange factor activity / response to amphetamine / G protein-coupled receptor binding ...Adenylate cyclase activating pathway / sensory perception of chemical stimulus / response to caffeine / G-protein alpha-subunit binding / Adenylate cyclase inhibitory pathway / adenylate cyclase regulator activity / protein folding chaperone / guanyl-nucleotide exchange factor activity / response to amphetamine / G protein-coupled receptor binding / G-protein beta/gamma-subunit complex binding / Olfactory Signaling Pathway / adenylate cyclase-activating G protein-coupled receptor signaling pathway / sensory perception of smell / adenylate cyclase-activating dopamine receptor signaling pathway / heterotrimeric G-protein complex / G protein activity / cell cortex / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / G protein-coupled receptor signaling pathway / GTPase activity / centrosome / GTP binding / signal transduction / extracellular exosome / metal ion binding / plasma membrane / cytoplasm / cytosol
Similarity search - Function
Synembryn / Guanine nucleotide exchange factor, Ric8 / Guanine nucleotide exchange factor synembryn / G-protein alpha subunit, group S / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / Armadillo-like helical ...Synembryn / Guanine nucleotide exchange factor, Ric8 / Guanine nucleotide exchange factor synembryn / G-protein alpha subunit, group S / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / Armadillo-like helical / Armadillo-type fold / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Guanine nucleotide-binding protein G(olf) subunit alpha / Chaperone Ric-8B
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsPapasergi-Scott, M.M. / Skiniotis, G.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01 GM088242 United States
CitationJournal: Structure / Year: 2023
Title: Structures of Ric-8B in complex with Gα protein folding clients reveal isoform specificity mechanisms.
Authors: Makaía M Papasergi-Scott / Frank E Kwarcinski / Maiya Yu / Ouliana Panova / Ann M Ovrutsky / Georgios Skiniotis / Gregory G Tall /
Abstract: Mammalian Ric-8 proteins act as chaperones to regulate the cellular abundance of heterotrimeric G protein α subunits. The Ric-8A isoform chaperones Gαi/o, Gα12/13, and Gαq/11 subunits, while Ric- ...Mammalian Ric-8 proteins act as chaperones to regulate the cellular abundance of heterotrimeric G protein α subunits. The Ric-8A isoform chaperones Gαi/o, Gα12/13, and Gαq/11 subunits, while Ric-8B acts on Gαs/olf subunits. Here, we determined cryoelectron microscopy (cryo-EM) structures of Ric-8B in complex with Gαs and Gαolf, revealing isoform differences in the relative positioning and contacts between the C-terminal α5 helix of Gα within the concave pocket formed by Ric-8 α-helical repeat elements. Despite the overall architectural similarity with our earlier structures of Ric-8A complexed to Gαq and Gαi1, Ric-8B distinctly accommodates an extended loop found only in Gαs/olf proteins. The structures, along with results from Ric-8 protein thermal stability assays and cell-based Gαolf folding assays, support a requirement for the Gα C-terminal region for binding specificity, and highlight that multiple structural elements impart specificity for Ric-8/G protein binding.
History
DepositionSep 23, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 1, 2023Provider: repository / Type: Initial release
Revision 1.1Apr 5, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2May 17, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Oct 9, 2024Group: Data collection / Refinement description / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Guanine nucleotide-binding protein G(olf) subunit alpha
B: Isoform 1 of Synembryn-B


Theoretical massNumber of molelcules
Total (without water)108,2982
Polymers108,2982
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, Negative staining transmission electron microscope, gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Guanine nucleotide-binding protein G(olf) subunit alpha / Adenylate cyclase-stimulating G alpha protein / olfactory type


Mass: 44370.430 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNAL / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P38405
#2: Protein Isoform 1 of Synembryn-B / Protein Ric-8B


Mass: 63928.043 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Ric8b / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q80XE1
Has ligand of interestN
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Ric-8B in complex with G protein subunit alpha olf / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 8
SpecimenConc.: 3.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid type: UltrAuFoil
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Calibrated magnification: 57050 X / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 2.49 sec. / Electron dose: 68.6 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 4670

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Processing

EM software
IDNameVersionCategory
1RELION3.1particle selection
2SerialEMimage acquisition
4RELION3.1CTF correction
9cryoSPARCinitial Euler assignment
10cryoSPARCfinal Euler assignment
11cryoSPARCclassification
12cryoSPARC3D reconstruction
13PHENIXmodel refinement
14Cootmodel refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 4695512
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 247416 / Symmetry type: POINT
Atomic model buildingB value: 171.1 / Protocol: AB INITIO MODEL / Space: REAL
Atomic model buildingPDB-ID: 8EL7

8el7


Accession code: 8EL7 / Source name: PDB / Type: experimental model

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