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- PDB-8tge: Crystal structure of the Methanosarcina mazei glutamine synthetas... -

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Basic information

Entry
Database: PDB / ID: 8tge
TitleCrystal structure of the Methanosarcina mazei glutamine synthetase in complex with GlnK1
Components
  • Glutamine synthetase
  • Nitrogen regulatory protein GlnK1
KeywordsLYASE / Glutamine synthetase / GlnK / PII / complex / dodecamer / trimer / T-loop
Function / homology
Function and homology information


glutamine synthetase / glutamine biosynthetic process / glutamine synthetase activity / regulation of nitrogen utilization / enzyme regulator activity / ATP binding / metal ion binding / cytoplasm
Similarity search - Function
Glutamine synthetase type I / Nitrogen regulatory protein P-II / Nitrogen regulatory protein P-II / Nitrogen regulatory protein PII / P-II protein family profile. / 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 type I / Nitrogen regulatory protein P-II / Nitrogen regulatory protein P-II / Nitrogen regulatory protein PII / P-II protein family profile. / 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 / Nitrogen regulatory PII-like, alpha/beta / Nitrogen regulatory protein PII/ATP phosphoribosyltransferase, C-terminal / Glutamine synthetase/guanido kinase, catalytic domain
Similarity search - Domain/homology
Glutamine synthetase / Nitrogen regulatory protein GlnK1
Similarity search - Component
Biological speciesMethanosarcina mazei Go1 (archaea)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.3 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: 2023
Title: M. mazei glutamine synthetase and glutamine synthetase-GlnK1 structures reveal enzyme regulation by oligomer modulation.
Authors: Maria A Schumacher / Raul Salinas / Brady A Travis / Rajiv Ranjan Singh / Nicholas Lent /
Abstract: Glutamine synthetases (GS) play central roles in cellular nitrogen assimilation. Although GS active-site formation requires the oligomerization of just two GS subunits, all GS form large, multi- ...Glutamine synthetases (GS) play central roles in cellular nitrogen assimilation. Although GS active-site formation requires the oligomerization of just two GS subunits, all GS form large, multi-oligomeric machines. Here we describe a structural dissection of the archaeal Methanosarcina mazei (Mm) GS and its regulation. We show that Mm GS forms unstable dodecamers. Strikingly, we show this Mm GS oligomerization property is leveraged for a unique mode of regulation whereby labile Mm GS hexamers are stabilized by binding the nitrogen regulatory protein, GlnK1. Our GS-GlnK1 structure shows that GlnK1 functions as molecular glue to affix GS hexamers together, stabilizing formation of GS active-sites. These data, therefore, reveal the structural basis for a unique form of enzyme regulation by oligomer modulation.
History
DepositionJul 12, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 15, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 29, 2023Group: 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

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Glutamine synthetase
B: Glutamine synthetase
D: Glutamine synthetase
Y: Glutamine synthetase
M: Glutamine synthetase
P: Glutamine synthetase
J: Nitrogen regulatory protein GlnK1
G: Nitrogen regulatory protein GlnK1
Z: Nitrogen regulatory protein GlnK1


Theoretical massNumber of molelcules
Total (without water)362,6929
Polymers362,6929
Non-polymers00
Water29,0761614
1
A: Glutamine synthetase
B: Glutamine synthetase
D: Glutamine synthetase
Y: Glutamine synthetase
M: Glutamine synthetase
P: Glutamine synthetase
J: Nitrogen regulatory protein GlnK1
G: Nitrogen regulatory protein GlnK1
Z: Nitrogen regulatory protein GlnK1

A: Glutamine synthetase
B: Glutamine synthetase
D: Glutamine synthetase
Y: Glutamine synthetase
M: Glutamine synthetase
P: Glutamine synthetase
J: Nitrogen regulatory protein GlnK1
G: Nitrogen regulatory protein GlnK1
Z: Nitrogen regulatory protein GlnK1


  • defined by author&software
  • 725 kDa, 18 polymers
Theoretical massNumber of molelcules
Total (without water)725,38418
Polymers725,38418
Non-polymers00
Water32418
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_555-x,y,-z1
Buried area74850 Å2
ΔGint-411 kcal/mol
Surface area208820 Å2
MethodPISA
Unit cell
Length a, b, c (Å)137.990, 178.030, 169.048
Angle α, β, γ (deg.)90.00, 90.36, 90.00
Int Tables number5
Space group name H-MI121

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Components

#1: Protein
Glutamine synthetase / / GS / Glutamate--ammonia ligase


Mass: 52827.926 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Methanosarcina mazei Go1 (archaea) / Gene: glnA1, MM_0964 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8PY99, glutamine synthetase
#2: Protein Nitrogen regulatory protein GlnK1


Mass: 15241.536 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Methanosarcina mazei Go1 (archaea) / Gene: glnK1, MM_0732 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8PYW7
#3: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 1614 / 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: 2.84 Å3/Da / Density % sol: 56.63 %
Crystal growTemperature: 273 K / Method: vapor diffusion, hanging drop / Details: PEG 400, 0.1 M MES pH 6.5

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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 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Apr 12, 2022
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 2.3→89.02 Å / Num. obs: 180295 / % possible obs: 99.6 % / Redundancy: 3.1 % / CC1/2: 0.994 / Rpim(I) all: 0.068 / Rsym value: 0.103 / Net I/σ(I): 8.3
Reflection shellResolution: 2.3→2.36 Å / Mean I/σ(I) obs: 1.8 / Num. unique obs: 13618 / CC1/2: 0.631 / Rpim(I) all: 0.447 / Rsym value: 0.618

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Processing

Software
NameVersionClassification
PHENIX(1.20.1_4487: ???)refinement
SCALAdata scaling
XDSdata reduction
BALBESphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.3→89.02 Å / SU ML: 0 / Cross valid method: THROUGHOUT / σ(F): 38.85 / Phase error: 21.57 / Stereochemistry target values: TWIN_LSQ_F
RfactorNum. reflection% reflection
Rfree0.223 1767 1 %
Rwork0.1647 --
obs0.3 180295 99.91 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.3→89.02 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms23170 0 0 1614 24784
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0086
X-RAY DIFFRACTIONf_angle_d0.936
X-RAY DIFFRACTIONf_dihedral_angle_d6.2783189
X-RAY DIFFRACTIONf_chiral_restr0.0533484
X-RAY DIFFRACTIONf_plane_restr0.0094185
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.3-2.360.29911250.29913621X-RAY DIFFRACTION99
2.36-2.430.32981410.307313720X-RAY DIFFRACTION99
2.43-2.510.30891400.303113706X-RAY DIFFRACTION99
2.51-2.60.30741290.302913710X-RAY DIFFRACTION99
2.6-2.70.36871430.317413716X-RAY DIFFRACTION99
2.7-2.830.34221300.314413708X-RAY DIFFRACTION99
2.83-2.980.34581320.316713683X-RAY DIFFRACTION99
2.98-3.160.36971380.327913708X-RAY DIFFRACTION99
3.16-3.410.35181360.339813753X-RAY DIFFRACTION99
3.41-3.750.3821340.335713767X-RAY DIFFRACTION99
3.75-4.290.35091390.310613780X-RAY DIFFRACTION99
4.29-5.410.28331400.275113771X-RAY DIFFRACTION99
Refinement TLS params.Method: refined / Origin x: -0.1103 Å / Origin y: -0.051 Å / Origin z: 27.1577 Å
111213212223313233
T0.21 Å2-0.0064 Å2-0.0018 Å2-0.2379 Å2-0.0021 Å2--0.1097 Å2
L0.1657 °2-0.032 °20.005 °2-0.2301 °2-0.0032 °2--0.1124 °2
S0.012 Å °-0.0798 Å °0.0007 Å °0.0763 Å °0.0024 Å °-0.0012 Å °0.0004 Å °0.0055 Å °-0.0146 Å °
Refinement TLS groupSelection details: all

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