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- PDB-3v00: Studies of a constitutively active G-alpha subunit provide insigh... -

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Basic information

Entry
Database: PDB / ID: 3v00
TitleStudies of a constitutively active G-alpha subunit provide insights into the mechanism of G protein activation.
ComponentsGuanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
KeywordsSIGNALING PROTEIN / GTPase / GTP binding / Transducer / SIGNAL TRANSDUCTION / CELL CYCLE
Function / homology
Function and homology information


Extra-nuclear estrogen signaling / Adenylate cyclase inhibitory pathway / negative regulation of cyclic-nucleotide phosphodiesterase activity / detection of light stimulus involved in visual perception / G protein-coupled receptor complex / Inactivation, recovery and regulation of the phototransduction cascade / Activation of the phototransduction cascade / negative regulation of synaptic transmission / GTPase activating protein binding / Adrenaline,noradrenaline inhibits insulin secretion ...Extra-nuclear estrogen signaling / Adenylate cyclase inhibitory pathway / negative regulation of cyclic-nucleotide phosphodiesterase activity / detection of light stimulus involved in visual perception / G protein-coupled receptor complex / Inactivation, recovery and regulation of the phototransduction cascade / Activation of the phototransduction cascade / negative regulation of synaptic transmission / GTPase activating protein binding / Adrenaline,noradrenaline inhibits insulin secretion / ADP signalling through P2Y purinoceptor 12 / G alpha (i) signalling events / neurotransmitter receptor localization to postsynaptic specialization membrane / photoreceptor outer segment membrane / G alpha (i) signalling events / acyl binding / phototransduction, visible light / response to light stimulus / phototransduction / positive regulation of protein localization to cell cortex / T cell migration / D2 dopamine receptor binding / response to prostaglandin E / G protein-coupled serotonin receptor binding / adenylate cyclase regulator activity / adenylate cyclase-inhibiting serotonin receptor signaling pathway / photoreceptor inner segment / cellular response to forskolin / regulation of mitotic spindle organization / positive regulation of cholesterol biosynthetic process / G protein-coupled receptor binding / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G-protein beta/gamma-subunit complex binding / photoreceptor disc membrane / GDP binding / heterotrimeric G-protein complex / G protein activity / midbody / cell cortex / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / postsynapse / G protein-coupled receptor signaling pathway / cell division / GTPase activity / centrosome / protein kinase binding / GTP binding / glutamatergic synapse / magnesium ion binding / protein-containing complex / metal ion binding / nucleus / plasma membrane / cytosol / cytoplasm
Similarity search - Function
GI Alpha 1, domain 2-like / GI Alpha 1, domain 2-like / G-protein alpha subunit, group I / 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 / P-loop containing nucleotide triphosphate hydrolases / Rossmann fold ...GI Alpha 1, domain 2-like / GI Alpha 1, domain 2-like / G-protein alpha subunit, group I / 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 / P-loop containing nucleotide triphosphate hydrolases / Rossmann fold / P-loop containing nucleoside triphosphate hydrolase / Orthogonal Bundle / 3-Layer(aba) Sandwich / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
GUANOSINE-5'-DIPHOSPHATE / Guanine nucleotide-binding protein G(t) subunit alpha-1 / Guanine nucleotide-binding protein G(i) subunit alpha-1
Similarity search - Component
Biological speciesBos taurus (domestic cattle)
Rattus norvegicus (Norway rat)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.9 Å
AuthorsSingh, G. / Cerione, R.A.
CitationJournal: Biochemistry / Year: 2012
Title: A constitutively active G-alpha subunit provide insights into the mechanism of G protein activation
Authors: Singh, G. / Ramachandran, S. / Cerione, R.A.
History
DepositionDec 7, 2011Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 18, 2012Provider: repository / Type: Initial release
Revision 1.1Jun 26, 2013Group: Database references
Revision 1.2Jul 26, 2017Group: Refinement description / Source and taxonomy / Category: entity_src_gen / software
Revision 1.3Sep 13, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_ref_seq_dif / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
A: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)124,3296
Polymers122,9993
Non-polymers1,3303
Water5,314295
1
C: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)41,4432
Polymers41,0001
Non-polymers4431
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)41,4432
Polymers41,0001
Non-polymers4431
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
A: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)41,4432
Polymers41,0001
Non-polymers4431
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
4
C: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
A: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules

C: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
A: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)248,65712
Polymers245,9986
Non-polymers2,6596
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation7_465y-1,x+1,-z1
Buried area24070 Å2
ΔGint-122 kcal/mol
Surface area97890 Å2
MethodPISA
5
C: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules

B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
A: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)124,3296
Polymers122,9993
Non-polymers1,3303
Water543
TypeNameSymmetry operationNumber
crystal symmetry operation7_465y-1,x+1,-z1
identity operation1_555x,y,z1
Buried area7920 Å2
ΔGint-49 kcal/mol
Surface area53060 Å2
MethodPISA
6
C: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules

A: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)82,8864
Polymers81,9992
Non-polymers8862
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation7_465y-1,x+1,-z1
Buried area4740 Å2
ΔGint-33 kcal/mol
Surface area36390 Å2
MethodPISA
7
B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules

B: Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)82,8864
Polymers81,9992
Non-polymers8862
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation7_465y-1,x+1,-z1
Buried area3950 Å2
ΔGint-23 kcal/mol
Surface area35730 Å2
MethodPISA
Unit cell
Length a, b, c (Å)93.173, 93.173, 380.580
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number96
Space group name H-MP43212

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Components

#1: Protein Guanine nucleotide-binding protein G(t) subunit alpha-1/ Guanine nucleotide-binding protein G(i) subunit alpha-1 chimeric protein / Transducin alpha-1 chain


Mass: 40999.656 Da / Num. of mol.: 3 / Fragment: UNP P04695 1-215 & 295-350, UNP P10824 220-298 / Mutation: G56P, K244H and D247N
Source method: isolated from a genetically manipulated source
Details: T7 LAC PROMOTER
Source: (gene. exp.) Bos taurus (domestic cattle), (gene. exp.) Rattus norvegicus (Norway rat)
Gene: GNAT1, GNAT1_BOVIN / Plasmid: PET VECTOR / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P04695, UniProt: P10824
#2: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Comment: GDP, energy-carrying molecule*YM
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 295 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.36 Å3/Da / Density % sol: 63.39 %
Crystal growTemperature: 295 K / Method: vapor diffusion, hanging drop / pH: 6.3
Details: 1.9 M Ammonium Sulphate in 0.05 M Sodium Cacodylate buffer, pH 6.3, VAPOR DIFFUSION, HANGING DROP, temperature 295K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: CHESS / Beamline: A1 / Wavelength: 0.978 Å
DetectorType: ADSC QUANTUM 210 / Detector: CCD / Date: Nov 8, 2007 / Details: Mirrors
RadiationMonochromator: Horizontal focusing 5.05 asymmetric cut Si(111)
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.978 Å / Relative weight: 1
ReflectionResolution: 2.7→39.74 Å / Num. all: 47386 / Num. obs: 47006 / % possible obs: 99.2 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 11.3 / Redundancy: 8.6 % / Biso Wilson estimate: 89.2 Å2 / Rmerge(I) obs: 0.08 / Rsym value: 0.08 / Net I/σ(I): 21.253
Reflection shellResolution: 2.7→2.8 Å / % possible all: 8.8

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Processing

Software
NameVersionClassification
CrystalCleardata collection
MOLREPphasing
CNS1.1refinement
DENZOdata reduction
HKL-2000data scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 1TAG

1tag


Resolution: 2.9→39.74 Å / Rfactor Rfree error: 0.005 / Cross valid method: THROUGHOUT / σ(F): 0 / Stereochemistry target values: ENGH & HUBER
RfactorNum. reflection% reflectionSelection details
Rfree0.272 3751 10 %RANDOM
Rwork0.221 ---
obs0.221 37430 97.4 %-
all-37430 --
Displacement parametersBiso mean: 94.5 Å2
Refine analyzeLuzzati coordinate error free: 0.46 Å / Luzzati sigma a free: 0.56 Å
Refinement stepCycle: LAST / Resolution: 2.9→39.74 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms8409 0 84 295 8788
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONc_bond_d0.007
X-RAY DIFFRACTIONc_bond_d_na
X-RAY DIFFRACTIONc_bond_d_prot
X-RAY DIFFRACTIONc_angle_d
X-RAY DIFFRACTIONc_angle_d_na
X-RAY DIFFRACTIONc_angle_d_prot
X-RAY DIFFRACTIONc_angle_deg1.1
X-RAY DIFFRACTIONc_angle_deg_na
X-RAY DIFFRACTIONc_angle_deg_prot
X-RAY DIFFRACTIONc_dihedral_angle_d23.3
X-RAY DIFFRACTIONc_dihedral_angle_d_na
X-RAY DIFFRACTIONc_dihedral_angle_d_prot
X-RAY DIFFRACTIONc_improper_angle_d0.81
X-RAY DIFFRACTIONc_improper_angle_d_na
X-RAY DIFFRACTIONc_improper_angle_d_prot
X-RAY DIFFRACTIONc_mcbond_it1.5
X-RAY DIFFRACTIONc_mcangle_it
X-RAY DIFFRACTIONc_scbond_it
X-RAY DIFFRACTIONc_scangle_it

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