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- PDB-7yzn: Structure of C-terminally truncated aIF5B from Pyrococcus abyssi ... -

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Basic information

Entry
Database: PDB / ID: 7yzn
TitleStructure of C-terminally truncated aIF5B from Pyrococcus abyssi complexed with GTP
ComponentsProbable translation initiation factor IF-2
KeywordsTRANSLATION / ribosome / initiator tRNA / initiation factor / eIF5B / IF2
Function / homology
Function and homology information


intein-mediated protein splicing / intron homing / translation initiation factor activity / endonuclease activity / GTPase activity / GTP binding / cytoplasm
Similarity search - Function
Translation initiation factor aIF-2, archaea / Elongation factor Tu-type domain / Elongation factor Tu domain 4 / Translation initiation factor IF- 2, domain 3 / Translation-initiation factor 2 / Translation initiation factor IF- 2 / Translation initiation factor IF-2, domain 3 superfamily / Intein splicing domain / Intein DOD homing endonuclease / Intein DOD-type homing endonuclease domain profile. ...Translation initiation factor aIF-2, archaea / Elongation factor Tu-type domain / Elongation factor Tu domain 4 / Translation initiation factor IF- 2, domain 3 / Translation-initiation factor 2 / Translation initiation factor IF- 2 / Translation initiation factor IF-2, domain 3 superfamily / Intein splicing domain / Intein DOD homing endonuclease / Intein DOD-type homing endonuclease domain profile. / Intein C-terminal splicing region / Intein C-terminal splicing motif profile. / Hint domain C-terminal / Hint (Hedgehog/Intein) domain C-terminal region / Intein N-terminal splicing region / Intein N-terminal splicing motif profile. / Hint domain N-terminal / Hint (Hedgehog/Intein) domain N-terminal region / Homing endonuclease / Hint domain superfamily / Translation elongation factor EFTu-like, domain 2 / Elongation factor Tu domain 2 / Translational (tr)-type GTP-binding domain / Elongation factor Tu GTP binding domain / Translational (tr)-type guanine nucleotide-binding (G) domain profile. / Small GTP-binding protein domain / Translation protein, beta-barrel domain superfamily / P-loop containing nucleotide triphosphate hydrolases / P-loop containing nucleoside triphosphate hydrolase / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
GUANOSINE-5'-TRIPHOSPHATE / Probable translation initiation factor IF-2
Similarity search - Component
Biological speciesPyrococcus abyssi GE5 (archaea)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.7 Å
AuthorsBourgeois, G. / Schmitt, E. / Mechulam, Y. / Coureux, P.D. / Kazan, R.
Funding support France, 2items
OrganizationGrant numberCountry
Centre National de la Recherche Scientifique (CNRS)UMR7654 France
Agence Nationale de la Recherche (ANR)ANR-17-CE11-0037 France
CitationJournal: Nucleic Acids Res / Year: 2022
Title: Role of aIF5B in archaeal translation initiation.
Authors: Ramy Kazan / Gabrielle Bourgeois / Christine Lazennec-Schurdevin / Eric Larquet / Yves Mechulam / Pierre-Damien Coureux / Emmanuelle Schmitt /
Abstract: In eukaryotes and in archaea late steps of translation initiation involve the two initiation factors e/aIF5B and e/aIF1A. In eukaryotes, the role of eIF5B in ribosomal subunit joining is established ...In eukaryotes and in archaea late steps of translation initiation involve the two initiation factors e/aIF5B and e/aIF1A. In eukaryotes, the role of eIF5B in ribosomal subunit joining is established and structural data showing eIF5B bound to the full ribosome were obtained. To achieve its function, eIF5B collaborates with eIF1A. However, structural data illustrating how these two factors interact on the small ribosomal subunit have long been awaited. The role of the archaeal counterparts, aIF5B and aIF1A, remains to be extensively addressed. Here, we study the late steps of Pyrococcus abyssi translation initiation. Using in vitro reconstituted initiation complexes and light scattering, we show that aIF5B bound to GTP accelerates subunit joining without the need for GTP hydrolysis. We report the crystallographic structures of aIF5B bound to GDP and GTP and analyze domain movements associated to these two nucleotide states. Finally, we present the cryo-EM structure of an initiation complex containing 30S bound to mRNA, Met-tRNAiMet, aIF5B and aIF1A at 2.7 Å resolution. Structural data shows how archaeal 5B and 1A factors cooperate to induce a conformation of the initiator tRNA favorable to subunit joining. Archaeal and eukaryotic features of late steps of translation initiation are discussed.
History
DepositionFeb 21, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 29, 2022Provider: repository / Type: Initial release
Revision 1.1Aug 31, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.2May 1, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Probable translation initiation factor IF-2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)53,6534
Polymers53,0821
Non-polymers5703
Water6,215345
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)72.260, 72.260, 205.820
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number92
Space group name H-MP41212
Space group name HallP4abw2nw
Symmetry operation#1: x,y,z
#2: -y+1/2,x+1/2,z+1/4
#3: y+1/2,-x+1/2,z+3/4
#4: x+1/2,-y+1/2,-z+3/4
#5: -x+1/2,y+1/2,-z+1/4
#6: -x,-y,z+1/2
#7: y,x,-z
#8: -y,-x,-z+1/2

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Components

#1: Protein Probable translation initiation factor IF-2


Mass: 53082.059 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus abyssi GE5 (archaea) / Strain: GE5 / Orsay / Gene: infB, PYRAB11390, PAB0755 / Plasmid: pET15blpa / Production host: Escherichia coli (E. coli) / References: UniProt: Q9UZK7
#2: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


Mass: 523.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: GTP, energy-carrying molecule*YM
#4: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Na
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 345 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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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: 2.53 Å3/Da / Density % sol: 51.4 %
Crystal growTemperature: 277 K / Method: vapor diffusion, sitting drop / pH: 5 / Details: 8% Tacsimate; 20% PEG3350

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SOLEIL / Beamline: PROXIMA 2 / Wavelength: 0.98 Å
DetectorType: DECTRIS EIGER X 9M / Detector: PIXEL / Date: Nov 20, 2020
RadiationMonochromator: CRYSTAL / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.98 Å / Relative weight: 1
ReflectionResolution: 1.7→50 Å / Num. obs: 61785 / % possible obs: 99.5 % / Redundancy: 26.3 % / Biso Wilson estimate: 27.37 Å2 / CC1/2: 0.999 / Rmerge(I) obs: 0.112 / Net I/σ(I): 20.2
Reflection shellResolution: 1.7→1.8 Å / Rmerge(I) obs: 1.885 / Mean I/σ(I) obs: 1.88 / Num. unique obs: 9577 / CC1/2: 0.712

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Processing

Software
NameVersionClassification
PHENIX1.18.2_3874refinement
XDSdata reduction
PHASERphasing
XSCALEdata scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: Pa-aIF5B

Resolution: 1.7→49.75 Å / SU ML: 0.2236 / Cross valid method: FREE R-VALUE / σ(F): 1.35 / Phase error: 18.7899
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.1964 2926 4.8 %
Rwork0.1689 58078 -
obs0.1702 61004 99.99 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 40.43 Å2
Refinement stepCycle: LAST / Resolution: 1.7→49.75 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3554 0 34 345 3933
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.01043732
X-RAY DIFFRACTIONf_angle_d1.04555082
X-RAY DIFFRACTIONf_chiral_restr0.3600
X-RAY DIFFRACTIONf_plane_restr0.0077664
X-RAY DIFFRACTIONf_dihedral_angle_d28.79571462
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
1.7-1.730.39661400.34592711X-RAY DIFFRACTION100
1.73-1.760.31761310.29382733X-RAY DIFFRACTION100
1.76-1.790.27241200.26342701X-RAY DIFFRACTION100
1.79-1.820.26551310.22552743X-RAY DIFFRACTION100
1.82-1.860.26821330.21322719X-RAY DIFFRACTION99.96
1.86-1.90.23791410.18662722X-RAY DIFFRACTION99.97
1.9-1.950.20231170.17622739X-RAY DIFFRACTION99.96
1.95-1.990.18371390.17472721X-RAY DIFFRACTION100
1.99-2.050.19191450.17512721X-RAY DIFFRACTION100
2.05-2.110.20011530.1712710X-RAY DIFFRACTION100
2.11-2.180.20611400.16262759X-RAY DIFFRACTION100
2.18-2.250.17471350.15682736X-RAY DIFFRACTION100
2.25-2.340.18061350.16232756X-RAY DIFFRACTION100
2.35-2.450.19881340.15742755X-RAY DIFFRACTION100
2.45-2.580.17941620.15482758X-RAY DIFFRACTION100
2.58-2.740.19171150.16122785X-RAY DIFFRACTION100
2.74-2.950.21721690.16832751X-RAY DIFFRACTION100
2.95-3.250.19451350.16932811X-RAY DIFFRACTION100
3.25-3.720.19331410.16032835X-RAY DIFFRACTION100
3.72-4.690.14831480.14082865X-RAY DIFFRACTION100
4.69-49.750.21151620.17823047X-RAY DIFFRACTION99.91
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
11.24180254774-0.0118400743505-0.1854936263850.3957184590030.028756345881.060354889270.00194909765507-0.0625860395047-0.0968917521002-0.0104198252543-0.0190746628082-0.02228050727960.0444313054716-0.003437314552330.0008936969260220.1040463262440.002581799871590.006606469759440.19549201081-0.0005516847604330.179103243922-4.2380660961-22.3372560446-22.5314547008
20.02940961764320.0306642826363-0.1387991928050.0131257488371-0.08007434153710.5535702226050.1273421497130.344473484370.125201917689-0.0755667255021-0.1182128368160.2385531200970.0923938487728-0.3475614429860.05563752279220.1843244233880.1139964419460.07274423848650.3832535163250.02489729882560.362820476562-13.1884655542-2.89700101503-38.4294642321
30.1346797684050.0171422903076-0.08083017057290.11850392607-0.09861324699050.1992564703840.03594318486950.2761827647380.508472555023-0.3435147204370.2099478525930.00822852002874-0.455369016266-0.1757340113920.0392347185390.5275272135150.02464969580670.1032659301620.2908600134550.08409285355660.5324798211072.145303012459.85116805338-30.5871419599
Refinement TLS group

Refine-ID: X-RAY DIFFRACTION / Auth asym-ID: A / Label asym-ID: A

IDRefine TLS-IDSelection detailsAuth seq-IDLabel seq-ID
11chain 'A' and (resid 5 through 331 )5 - 3313 - 351
22chain 'A' and (resid 332 through 364 )332 - 364352 - 386
33chain 'A' and (resid 365 through 455 )365 - 455387 - 484

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