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- PDB-7tf9: L. monocytogenes GS(14)-Q-GlnR peptide -

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Basic information

Entry
Database: PDB / ID: 7tf9
TitleL. monocytogenes GS(14)-Q-GlnR peptide
Components
  • C-tail peptide of Glutamine synthetase repressor
  • Glutamine synthetase
KeywordsBIOSYNTHETIC PROTEIN / LIGASE / glutamine synthetase repressor tetradecamer
Function / homology
Function and homology information


glutamine synthetase / glutamine biosynthetic process / glutamine synthetase activity / regulation of DNA-templated transcription / DNA binding / ATP binding / metal ion binding / cytoplasm
Similarity search - Function
MerR-type HTH domain signature. / Glutamine synthetase type I / Glutamine synthetase, N-terminal conserved site / MerR HTH family regulatory protein / Glutamine synthetase signature 1. / Glutamine synthetase, beta-Grasp domain / helix_turn_helix, mercury resistance / MerR-type HTH domain profile. / MerR-type HTH domain / Glutamine synthetase (GS) beta-grasp domain profile. ...MerR-type HTH domain signature. / Glutamine synthetase type I / Glutamine synthetase, N-terminal conserved site / MerR HTH family regulatory protein / Glutamine synthetase signature 1. / Glutamine synthetase, beta-Grasp domain / helix_turn_helix, mercury resistance / MerR-type HTH domain profile. / MerR-type HTH domain / Glutamine synthetase (GS) beta-grasp domain profile. / Glutamine synthetase, glycine-rich site / Glutamine synthetase putative ATP-binding region signature. / Glutamine synthetase, N-terminal domain superfamily / Glutamine synthetase (GS) catalytic domain profile. / Glutamine synthetase, catalytic domain / Glutamine synthetase, N-terminal domain / Glutamine synthetase, catalytic domain / Glutamine synthetase, catalytic domain / Glutamine synthetase/guanido kinase, catalytic domain / Putative DNA-binding domain superfamily
Similarity search - Domain/homology
GLUTAMINE / Glutamine synthetase / Glutamine synthetase repressor
Similarity search - Component
Biological speciesListeria monocytogenes (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.61 Å
AuthorsTravis, B.A. / Peck, J. / Schumacher, M.A.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35-GM130290 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)F31-AI150138 United States
National Institutes of Health/National Institute of Environmental Health Sciences (NIH/NIEHS)ZIC-ES103326 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 6, 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.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2Feb 28, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Glutamine synthetase
B: Glutamine synthetase
C: Glutamine synthetase
D: C-tail peptide of Glutamine synthetase repressor
E: C-tail peptide of Glutamine synthetase repressor
F: Glutamine synthetase
G: Glutamine synthetase
H: Glutamine synthetase
I: Glutamine synthetase
J: C-tail peptide of Glutamine synthetase repressor
K: C-tail peptide of Glutamine synthetase repressor
L: C-tail peptide of Glutamine synthetase repressor
M: C-tail peptide of Glutamine synthetase repressor
N: Glutamine synthetase
O: Glutamine synthetase
P: Glutamine synthetase
Q: Glutamine synthetase
R: Glutamine synthetase
S: Glutamine synthetase
T: Glutamine synthetase
U: C-tail peptide of Glutamine synthetase repressor
V: C-tail peptide of Glutamine synthetase repressor
W: C-tail peptide of Glutamine synthetase repressor
X: C-tail peptide of Glutamine synthetase repressor
Y: C-tail peptide of Glutamine synthetase repressor
Z: C-tail peptide of Glutamine synthetase repressor
a: C-tail peptide of Glutamine synthetase repressor
b: C-tail peptide of Glutamine synthetase repressor
hetero molecules


Theoretical massNumber of molelcules
Total (without water)750,67070
Polymers747,94428
Non-polymers2,72742
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Glutamine synthetase


Mass: 52763.766 Da / Num. of mol.: 14
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: Protein/peptide
C-tail peptide of Glutamine synthetase repressor


Mass: 660.784 Da / Num. of mol.: 14 / Fragment: Residues 118-122 / Source method: obtained synthetically / Source: (synth.) Listeria monocytogenes (bacteria) / References: UniProt: A0A807UZD6
#3: Chemical...
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 28 / Source method: obtained synthetically / Formula: Mg
#4: Chemical
ChemComp-GLN / GLUTAMINE


Type: L-peptide linking / Mass: 146.144 Da / Num. of mol.: 14 / Source method: obtained synthetically / Formula: C5H10N2O3
Has ligand of interestN

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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: Tetradecameric L. monocytogenes GS complex with glutamine and GlnR C-tail peptides
Type: COMPLEX / Entity ID: #1-#2 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Buffer solutionpH: 7.5
SpecimenConc.: 0.9 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2200 nm / Nominal defocus min: 300 nm
Image recordingElectron dose: 42.65 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameVersionCategory
2SerialEMimage acquisition
4cryoSPARC3.2CTF correction
7Coot0.9model fitting
9PHENIX1.19model refinement
10cryoSPARC3.2initial Euler assignment
11cryoSPARC3.2final Euler assignment
12cryoSPARC3.2classification
13cryoSPARC3.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: D7 (2x7 fold dihedral)
3D reconstructionResolution: 2.61 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 212272 / Symmetry type: POINT
Atomic model buildingSpace: REAL
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 107.63 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.004750918
ELECTRON MICROSCOPYf_angle_d0.682668936
ELECTRON MICROSCOPYf_chiral_restr0.05027462
ELECTRON MICROSCOPYf_plane_restr0.00429030
ELECTRON MICROSCOPYf_dihedral_angle_d15.756518788

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