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- PDB-7ten: Crystal structure of the Listeria monocytogenes GS-Met-Sox-P- ADP... -

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Basic information

Entry
Database: PDB / ID: 7ten
TitleCrystal structure of the Listeria monocytogenes GS-Met-Sox-P- ADP complex to 3.5 Angstrom
ComponentsGlutamine synthetase
KeywordsLIGASE/INHIBITOR / glutamate ammonium ligase / GlnR / GS / glutamine synthetase / Met-Sox / transition state / LIGASE / LIGASE-INHIBITOR complex
Function / homology
Function and homology information


glutamine synthetase / glutamine biosynthetic process / glutamine synthetase activity / ATP binding / metal ion binding / cytoplasm
Similarity search - Function
Glutamine synthetase type I / Glutamine synthetase (GS) beta-grasp domain profile. / Glutamine synthetase (GS) catalytic domain profile. / Glutamine synthetase, N-terminal conserved site / Glutamine synthetase signature 1. / Glutamine synthetase, beta-Grasp domain / Glutamine synthetase, glycine-rich site / Glutamine synthetase putative ATP-binding region signature. / Glutamine synthetase, N-terminal domain / Glutamine synthetase, N-terminal domain superfamily ...Glutamine synthetase type I / Glutamine synthetase (GS) beta-grasp domain profile. / Glutamine synthetase (GS) catalytic domain profile. / Glutamine synthetase, N-terminal conserved site / Glutamine synthetase signature 1. / Glutamine synthetase, beta-Grasp domain / Glutamine synthetase, glycine-rich site / Glutamine synthetase putative ATP-binding region signature. / Glutamine synthetase, N-terminal domain / Glutamine synthetase, N-terminal domain superfamily / Glutamine synthetase, catalytic domain / Glutamine synthetase, catalytic domain / Glutamine synthetase, catalytic domain / Glutamine synthetase/guanido kinase, catalytic domain
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / L-METHIONINE-S-SULFOXIMINE PHOSPHATE / Glutamine synthetase
Similarity search - Component
Biological speciesListeria monocytogenes (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 3.5 Å
AuthorsSchumacher, M.A.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35-GM130290 United States
CitationJournal: Nat Commun / Year: 2022
Title: Molecular dissection of the glutamine synthetase-GlnR nitrogen regulatory circuitry in Gram-positive bacteria.
Authors: Brady A Travis / Jared V Peck / Raul Salinas / Brandon Dopkins / Nicholas Lent / Viet D Nguyen / Mario J Borgnia / Richard G Brennan / Maria A Schumacher /
Abstract: How bacteria sense and respond to nitrogen levels are central questions in microbial physiology. In Gram-positive bacteria, nitrogen homeostasis is controlled by an operon encoding glutamine ...How bacteria sense and respond to nitrogen levels are central questions in microbial physiology. In Gram-positive bacteria, nitrogen homeostasis is controlled by an operon encoding glutamine synthetase (GS), a dodecameric machine that assimilates ammonium into glutamine, and the GlnR repressor. GlnR detects nitrogen excess indirectly by binding glutamine-feedback-inhibited-GS (FBI-GS), which activates its transcription-repression function. The molecular mechanisms behind this regulatory circuitry, however, are unknown. Here we describe biochemical and structural analyses of GS and FBI-GS-GlnR complexes from pathogenic and non-pathogenic Gram-positive bacteria. The structures show FBI-GS binds the GlnR C-terminal domain within its active-site cavity, juxtaposing two GlnR monomers to form a DNA-binding-competent GlnR dimer. The FBI-GS-GlnR interaction stabilizes the inactive GS conformation. Strikingly, this interaction also favors a remarkable dodecamer to tetradecamer transition in some GS, breaking the paradigm that all bacterial GS are dodecamers. These data thus unveil unique structural mechanisms of transcription and enzymatic regulation.
History
DepositionJan 5, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 29, 2022Provider: repository / Type: Initial release
Revision 1.1Aug 10, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Oct 18, 2023Group: 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

Downloads & links

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Assembly

Deposited unit
A: Glutamine synthetase
E: Glutamine synthetase
B: Glutamine synthetase
D: Glutamine synthetase
H: Glutamine synthetase
J: Glutamine synthetase
M: Glutamine synthetase
O: Glutamine synthetase
Q: Glutamine synthetase
S: Glutamine synthetase
U: Glutamine synthetase
W: Glutamine synthetase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)618,74536
Polymers610,49612
Non-polymers8,24924
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area76680 Å2
ΔGint-409 kcal/mol
Surface area182170 Å2
MethodPISA
Unit cell
Length a, b, c (Å)112.608, 137.613, 138.023
Angle α, β, γ (deg.)60.750, 87.160, 68.400
Int Tables number1
Space group name H-MP1

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Components

#1: Protein
Glutamine synthetase /


Mass: 50874.691 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Listeria monocytogenes (bacteria)
Gene: glnA, BW273_06565, CW834_10155, E3W32_09805, E5H26_02065, FL871_07615, GF092_00575, GIG92_01785, GIH49_01255, HF764_002412
Production host: Escherichia coli (E. coli) / References: UniProt: A0A5D5GA79, glutamine synthetase
#2: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate


Mass: 427.201 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
#3: Chemical
ChemComp-P3S / L-METHIONINE-S-SULFOXIMINE PHOSPHATE


Mass: 260.205 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C5H13N2O6PS / Feature type: SUBJECT OF INVESTIGATION
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: 2.81 Å3/Da / Density % sol: 56.15 %
Crystal growTemperature: 298 K / Method: vapor diffusion, hanging drop
Details: 15% PEG 4000, 40 mM potassium phosphate dibasic pH 7.5, 20% glycerol

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: ALS / Beamline: 5.0.2 / Wavelength: 1.01 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Mar 12, 2021
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.01 Å / Relative weight: 1
ReflectionResolution: 3.5→63.88 Å / Num. obs: 62204 / % possible obs: 74.7 % / Redundancy: 3.1 % / CC1/2: 0.995 / Rpim(I) all: 0.13 / Rsym value: 0.184 / Net I/σ(I): 4.8
Reflection shellResolution: 3.5→3.62 Å / Mean I/σ(I) obs: 1.2 / Num. unique obs: 6451 / CC1/2: 0.323 / Rpim(I) all: 0.966 / Rsym value: 1.15

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Phasing

PhasingMethod: molecular replacement

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Processing

Software
NameVersionClassificationNB
XDSdata reduction
SCALAdata scaling
PHASERphasing
PHENIX1.17.1_3660refinement
PDB_EXTRACT3.27data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 7TDP

7tdp


Resolution: 3.5→63.88 Å / SU ML: 0.55 / Cross valid method: THROUGHOUT / σ(F): 1.96 / Phase error: 30.51 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2726 1988 3.2 %
Rwork0.2013 60215 -
obs0.2035 62203 74.33 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 283.39 Å2 / Biso mean: 113.2813 Å2 / Biso min: 42.11 Å2
Refinement stepCycle: final / Resolution: 3.5→63.88 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms42079 0 504 0 42583
Biso mean--108.61 --
Num. residues----5304
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00543620
X-RAY DIFFRACTIONf_angle_d0.95259144
X-RAY DIFFRACTIONf_dihedral_angle_d19.1175981
X-RAY DIFFRACTIONf_chiral_restr0.0526368
X-RAY DIFFRACTIONf_plane_restr0.0077692
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 14

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
3.5-3.590.35261550.31214469462477
3.59-3.680.3761530.31354392454576
3.68-3.790.34421430.30634347449075
3.79-3.920.33891270.28754292441974
3.92-4.060.30671480.27594313446175
4.06-4.220.33511500.24814378452876
4.22-4.410.3231400.21744347448775
4.41-4.640.27961540.19524259441374
4.64-4.930.24471350.18144226436173
4.93-5.310.23121400.17534300444074
5.31-5.850.29041360.19294271440774
5.85-6.690.271320.19274247437973
6.69-8.430.24571400.18044257439774
8.43-63.880.19571350.11744117425271
Refinement TLS params.Method: refined / Origin x: 10.124 Å / Origin y: -44.8282 Å / Origin z: 0.4931 Å
111213212223313233
T0.4726 Å2-0.0208 Å2-0.0129 Å2-0.4834 Å20.0189 Å2--0.5158 Å2
L0.7068 °2-0.1305 °20.0633 °2-0.7085 °20.1592 °2--0.6703 °2
S0.103 Å °0.027 Å °-0.1017 Å °-0.1116 Å °-0.0209 Å °-0.0959 Å °0.0886 Å °-0.0568 Å °-0.0739 Å °
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1allA3 - 444
2X-RAY DIFFRACTION1allF501 - 601
3X-RAY DIFFRACTION1allE3 - 444
4X-RAY DIFFRACTION1allK501 - 601
5X-RAY DIFFRACTION1allB3 - 444
6X-RAY DIFFRACTION1allC501 - 601
7X-RAY DIFFRACTION1allD3 - 444
8X-RAY DIFFRACTION1allG501 - 601
9X-RAY DIFFRACTION1allH3 - 444
10X-RAY DIFFRACTION1allI501 - 601
11X-RAY DIFFRACTION1allJ3 - 444
12X-RAY DIFFRACTION1allL501 - 601
13X-RAY DIFFRACTION1allM3 - 444
14X-RAY DIFFRACTION1allN501 - 601
15X-RAY DIFFRACTION1allO3 - 444
16X-RAY DIFFRACTION1allP501 - 601
17X-RAY DIFFRACTION1allQ3 - 444
18X-RAY DIFFRACTION1allR501 - 601
19X-RAY DIFFRACTION1allS3 - 444
20X-RAY DIFFRACTION1allT501 - 601
21X-RAY DIFFRACTION1allU3 - 444
22X-RAY DIFFRACTION1allV501 - 601
23X-RAY DIFFRACTION1allW3 - 444
24X-RAY DIFFRACTION1allX501 - 601

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