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- PDB-6u9h: Arabidopsis thaliana acetohydroxyacid synthase complex -

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Basic information

Entry
Database: PDB / ID: 6u9h
TitleArabidopsis thaliana acetohydroxyacid synthase complex
Components
  • Acetolactate synthase small subunit 2, chloroplastic
  • Acetolactate synthase, chloroplastic
KeywordsPLANT PROTEIN / Complex
Function / homology
Function and homology information


acetolactate synthase regulator activity / acetolactate synthase activity / acetolactate synthase / regulation of catalytic activity / valine biosynthetic process / isoleucine biosynthetic process / response to herbicide / thiamine pyrophosphate binding / chloroplast stroma / chloroplast ...acetolactate synthase regulator activity / acetolactate synthase activity / acetolactate synthase / regulation of catalytic activity / valine biosynthetic process / isoleucine biosynthetic process / response to herbicide / thiamine pyrophosphate binding / chloroplast stroma / chloroplast / peroxisome / : / flavin adenine dinucleotide binding / response to hypoxia / magnesium ion binding
Similarity search - Function
ACT domain / Acetolactate synthase, small subunit, C-terminal / Small subunit of acetolactate synthase / Acetolactate synthase, small subunit / AHAS, ACT domain / ACT-like. Chain A, domain 2 / Acetolactate synthase/Transcription factor NikR, C-terminal / Acetolactate synthase, large subunit, biosynthetic / Acetolactate synthase large subunit, TPP binding domain / ACT domain ...ACT domain / Acetolactate synthase, small subunit, C-terminal / Small subunit of acetolactate synthase / Acetolactate synthase, small subunit / AHAS, ACT domain / ACT-like. Chain A, domain 2 / Acetolactate synthase/Transcription factor NikR, C-terminal / Acetolactate synthase, large subunit, biosynthetic / Acetolactate synthase large subunit, TPP binding domain / ACT domain / ACT domain / Thiamine pyrophosphate enzyme / TPP-binding enzyme, conserved site / Thiamine pyrophosphate enzymes signature. / ACT domain profile. / ACT domain / Thiamine pyrophosphate enzyme, central domain / Thiamine pyrophosphate enzyme, central domain / ACT-like domain / Thiamine pyrophosphate enzyme, N-terminal TPP-binding domain / Thiamine pyrophosphate enzyme, N-terminal TPP binding domain / Thiamine pyrophosphate enzyme, C-terminal TPP-binding / Thiamine pyrophosphate enzyme, C-terminal TPP binding domain / Thiamin diphosphate (ThDP)-binding fold, Pyr/PP domains / TPP-binding domain / Thiamin diphosphate-binding fold / DHS-like NAD/FAD-binding domain superfamily / Alpha-Beta Plaits / Rossmann fold / 2-Layer Sandwich / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
FLAVIN-ADENINE DINUCLEOTIDE / THIAMINE DIPHOSPHATE / Acetolactate synthase, chloroplastic / Acetolactate synthase small subunit 1, chloroplastic
Similarity search - Component
Biological speciesArabidopsis thaliana (thale cress)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsGuddat, L.W. / Low, Y.S. / Rao, Z.
Funding support Australia, 2items
OrganizationGrant numberCountry
National Health and Medical Research Council (NHMRC, Australia)1087713 Australia
National Health and Medical Research Council (NHMRC, Australia)1147297 Australia
CitationJournal: Nature / Year: 2020
Title: Structures of fungal and plant acetohydroxyacid synthases.
Authors: Thierry Lonhienne / Yu Shang Low / Mario D Garcia / Tristan Croll / Yan Gao / Quan Wang / Lou Brillault / Craig M Williams / James A Fraser / Ross P McGeary / Nicholas P West / Michael J ...Authors: Thierry Lonhienne / Yu Shang Low / Mario D Garcia / Tristan Croll / Yan Gao / Quan Wang / Lou Brillault / Craig M Williams / James A Fraser / Ross P McGeary / Nicholas P West / Michael J Landsberg / Zihe Rao / Gerhard Schenk / Luke W Guddat /
Abstract: Acetohydroxyacid synthase (AHAS), also known as acetolactate synthase, is a flavin adenine dinucleotide-, thiamine diphosphate- and magnesium-dependent enzyme that catalyses the first step in the ...Acetohydroxyacid synthase (AHAS), also known as acetolactate synthase, is a flavin adenine dinucleotide-, thiamine diphosphate- and magnesium-dependent enzyme that catalyses the first step in the biosynthesis of branched-chain amino acids. It is the target for more than 50 commercial herbicides. AHAS requires both catalytic and regulatory subunits for maximal activity and functionality. Here we describe structures of the hexadecameric AHAS complexes of Saccharomyces cerevisiae and dodecameric AHAS complexes of Arabidopsis thaliana. We found that the regulatory subunits of these AHAS complexes form a core to which the catalytic subunit dimers are attached, adopting the shape of a Maltese cross. The structures show how the catalytic and regulatory subunits communicate with each other to provide a pathway for activation and for feedback inhibition by branched-chain amino acids. We also show that the AHAS complex of Mycobacterium tuberculosis adopts a similar structure, thus demonstrating that the overall AHAS architecture is conserved across kingdoms.
History
DepositionSep 8, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 15, 2020Provider: repository / Type: Initial release
Revision 1.1Jan 27, 2021Group: Database references / Derived calculations / Category: citation / citation_author / struct_conn
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _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 / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id
Revision 1.2Mar 20, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

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  • Deposited structure unit
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Structure viewerMolecule:
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Assembly

Deposited unit
A: Acetolactate synthase, chloroplastic
B: Acetolactate synthase, chloroplastic
F: Acetolactate synthase small subunit 2, chloroplastic
G: Acetolactate synthase small subunit 2, chloroplastic
H: Acetolactate synthase, chloroplastic
I: Acetolactate synthase, chloroplastic
J: Acetolactate synthase small subunit 2, chloroplastic
K: Acetolactate synthase small subunit 2, chloroplastic
L: Acetolactate synthase, chloroplastic
M: Acetolactate synthase, chloroplastic
N: Acetolactate synthase small subunit 2, chloroplastic
O: Acetolactate synthase small subunit 2, chloroplastic
P: Acetolactate synthase, chloroplastic
Q: Acetolactate synthase, chloroplastic
R: Acetolactate synthase small subunit 2, chloroplastic
S: Acetolactate synthase small subunit 2, chloroplastic
hetero molecules


Theoretical massNumber of molelcules
Total (without water)953,67440
Polymers943,79316
Non-polymers9,88124
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: cross-linking
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Acetolactate synthase, chloroplastic / / AtALS / Acetohydroxy-acid synthase / Protein CHLORSULFURON RESISTANT 1


Mass: 64025.203 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: ALS, AHAS, CSR1, TZP5, At3g48560, T8P19.70 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: P17597, acetolactate synthase
#2: Protein
Acetolactate synthase small subunit 2, chloroplastic / / Acetohydroxy-acid synthase small subunit / ALS


Mass: 53948.906 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: At2g31810, F20M17.15 / Production host: Escherichia coli (E. coli) / References: UniProt: Q93YZ7
#3: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Mg
#4: Chemical
ChemComp-FAD / FLAVIN-ADENINE DINUCLEOTIDE / Flavin adenine dinucleotide


Mass: 785.550 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C27H33N9O15P2 / Comment: FAD*YM
#5: Chemical
ChemComp-TPP / THIAMINE DIPHOSPHATE / Thiamine pyrophosphate


Mass: 425.314 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C12H19N4O7P2S
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: Acetohydroxyacid synthase quaternary complex / Type: COMPLEX / Details: Recombinant expression from E. coli. / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 0.72 MDa / Experimental value: NO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenConc.: 0.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Glutaraldehyde cross linked
Specimen supportGrid material: COPPER
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELDBright-field microscopy
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3.2 sec. / Electron dose: 60 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 2

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.14_3260refinement
PHENIX1.14_3260refinement
EM software
IDNameVersionCategory
1Gautomatchparticle selection
4GctfCTF correction
11RELION3classification
12RELION33D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 76 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
15K6Q

5k6q

15K6Q1PDBexperimental model
22PC6

2pc6

12PC62PDBexperimental model
RefinementStereochemistry target values: CDL v1.2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.008547284
ELECTRON MICROSCOPYf_angle_d1.086164288
ELECTRON MICROSCOPYf_chiral_restr0.06597316
ELECTRON MICROSCOPYf_plane_restr0.00828328
ELECTRON MICROSCOPYf_dihedral_angle_d9.838736808

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