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- PDB-7tec: Structure of the Listeria monocytogenes GlnR-DNA complex to 3.45 ... -

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Basic information

Entry
Database: PDB / ID: 7tec
TitleStructure of the Listeria monocytogenes GlnR-DNA complex to 3.45 Angstrom
Components
  • DNA (5'-D(*CP*GP*TP*GP*TP*CP*AP*GP*AP*TP*AP*AP*TP*CP*TP*GP*AP*CP*AP*CP*G)-3')
  • HTH-type transcriptional regulator GlnR
KeywordsDNA BINDING PROTEIN/DNA / GlnR / listeria / glutamine synthetase / repressor / transcription / glnRA / DNA BINDING PROTEIN / DNA BINDING PROTEIN-DNA complex
Function / homology
Function and homology information


regulation of DNA-templated transcription / DNA binding
Similarity search - Function
MerR-type HTH domain signature. / MerR HTH family regulatory protein / MerR-type HTH domain profile. / helix_turn_helix, mercury resistance / MerR-type HTH domain / Putative DNA-binding domain superfamily
Similarity search - Domain/homology
DNA / DNA (> 10) / HTH-type transcriptional regulator GlnR
Similarity search - Component
Biological speciesListeria monocytogenes (bacteria)
synthetic construct (others)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 3.45 Å
AuthorsSchumacher, M.A. / Brennan, R.G.
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 4, 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

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Assembly

Deposited unit
A: HTH-type transcriptional regulator GlnR
H: DNA (5'-D(*CP*GP*TP*GP*TP*CP*AP*GP*AP*TP*AP*AP*TP*CP*TP*GP*AP*CP*AP*CP*G)-3')


Theoretical massNumber of molelcules
Total (without water)21,0862
Polymers21,0862
Non-polymers00
Water00
1
A: HTH-type transcriptional regulator GlnR
H: DNA (5'-D(*CP*GP*TP*GP*TP*CP*AP*GP*AP*TP*AP*AP*TP*CP*TP*GP*AP*CP*AP*CP*G)-3')

A: HTH-type transcriptional regulator GlnR
H: DNA (5'-D(*CP*GP*TP*GP*TP*CP*AP*GP*AP*TP*AP*AP*TP*CP*TP*GP*AP*CP*AP*CP*G)-3')


Theoretical massNumber of molelcules
Total (without water)42,1724
Polymers42,1724
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation3_555-x,y,-z+1/21
Buried area5300 Å2
ΔGint-33 kcal/mol
Surface area14460 Å2
MethodPISA
Unit cell
Length a, b, c (Å)28.632, 47.717, 121.410
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number17
Space group name H-MP2221

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Components

#1: Protein HTH-type transcriptional regulator GlnR / MerR family transcriptional regulator


Mass: 14638.857 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Listeria monocytogenes (bacteria)
Gene: glnR, A8L61_07010, ABZ57_05085, AF006_07180, AF817_01990, AMC55_04605, APD94_02400, ART25_07755, ARV28_05490, ARV77_10415, B1N06_02425, B1N38_06025, B1N40_09865, B1N52_08095, B1N70_08585, B1N71_ ...Gene: glnR, A8L61_07010, ABZ57_05085, AF006_07180, AF817_01990, AMC55_04605, APD94_02400, ART25_07755, ARV28_05490, ARV77_10415, B1N06_02425, B1N38_06025, B1N40_09865, B1N52_08095, B1N70_08585, B1N71_01820, B1O10_02055, B1O25_08310, B2H14_10505, B4X79_13085, B4Y29_01290, B4Y40_11845, B4Y47_05380, B4Y49_02050, B4Y56_02055, B4Y57_02055, B5K54_13065, B6112_07615, B6O07_10740, BB997_08390, BCZ19_06080, BW273_06570, C6S26_02850, CW834_10150, CW845_07630, CW895_11495, CX098_14230, D4164_01995, D4271_02955, D4900_04675, D4920_02385, D4947_02360, D4C60_03330, D4D22_02955, D4D89_02780, D4U00_01965, D4U23_10015, D5M63_02050, D5N24_04355, D6P18_02820, D7104_08235, DCK28_08790, DCT16_08455, E0I39_02830, E1V33_00335, E1W43_05380, E1W56_05615, E3W32_09810, E5F58_10230, E5H26_02070, EPC87_01440, EX365_07695, EXZ73_06750, EYY39_05995, F3O35_05110, F3R75_05795, F6436_09515, F6515_11560, FA835_14895, FC284_14445, FJU19_02820, FL871_07620, FLQ97_14195, FLR03_05055, FLR11_13675, FNX31_12240, FNX40_10035, FORC68_1320, FR217_04555, FV747_02470, G3O21_002452, G3R95_000630, GEK29_02375, GFK29_08620, GH165_00575, GHH22_03975, GHM39_05980, GIG92_01790, GIH49_01260, GJW51_07325, GNG71_06855, GON91_01810, GT011_02015, GXB45_00580, GYN46_14485, GYO86_08555, GYP27_02015, GYS19_13395, GYU05_01010, GYU24_02035, GYX95_04495, GYY14_11060, GYZ23_12950, GYZ33_11570, GYZ61_07200, GZI09_15320, GZK40_07615, GZM52_07550, GZN68_09475, HP506_001182, HQN34_002361, I6I37_04555, I8J47_00192, IP987_001477, JKV73_01561, JKV74_07850, JKV76_01310, JKV77_01646, JKX60_08595, JKX61_06825, KV70_04590, KW30_04605, LmNIHS28_00654, M643_00970, QD52_08715, R019_12835, UI29_08755, UP23_04390
Production host: Escherichia coli (E. coli) / References: UniProt: L8DSZ4
#2: DNA chain DNA (5'-D(*CP*GP*TP*GP*TP*CP*AP*GP*AP*TP*AP*AP*TP*CP*TP*GP*AP*CP*AP*CP*G)-3')


Mass: 6447.185 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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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: 1.97 Å3/Da / Density % sol: 37.54 %
Crystal growTemperature: 275 K / Method: vapor diffusion, hanging drop
Details: 50 mM sodium cacodylate pH 6.5, 35% MPD, 10 mM MgCl2

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: ALS / Beamline: 5.0.2 / Wavelength: 1.09 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Feb 12, 2021
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.09 Å / Relative weight: 1
ReflectionResolution: 3.45→37.51 Å / Num. obs: 2384 / % possible obs: 96.8 % / Redundancy: 4.1 % / CC1/2: 0.998 / Rpim(I) all: 0.038 / Rsym value: 0.064 / Net I/σ(I): 8.9
Reflection shellResolution: 3.45→4.34 Å / Mean I/σ(I) obs: 2 / Num. unique obs: 237 / CC1/2: 0.71 / Rpim(I) all: 0.24 / Rsym value: 0.42

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Phasing

PhasingMethod: molecular replacement

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Processing

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

7tea


Resolution: 3.45→37.51 Å / SU ML: 0.34 / Cross valid method: THROUGHOUT / σ(F): 1.36 / Phase error: 23.48 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.3091 238 9.98 %
Rwork0.2656 2146 -
obs0.2701 2384 96.79 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 165.47 Å2 / Biso mean: 64.3464 Å2 / Biso min: 20 Å2
Refinement stepCycle: final / Resolution: 3.45→37.51 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms548 428 0 0 976
Num. residues----93
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 2 / % reflection obs: 97 %

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all
3.45-4.340.31741140.252110361150
4.35-37.510.30471240.273511101234
Refinement TLS params.Method: refined / Origin x: 2.038 Å / Origin y: 5.37 Å / Origin z: 23.3464 Å
111213212223313233
T0.7511 Å2-0.1205 Å2-0.0752 Å2-0.4795 Å2-0.0169 Å2--0.2185 Å2
L1.907 °2-1.5792 °2-0.1568 °2-2.0865 °20.2662 °2--4.3892 °2
S0.2381 Å °0.2669 Å °-0.0324 Å °-0.4476 Å °0.0002 Å °-0.2037 Å °-0.0605 Å °0.5488 Å °-0.1829 Å °
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1allA2 - 73
2X-RAY DIFFRACTION1allH0 - 20

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