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- PDB-6mgt: Crystal structure of alpha-Amino-beta-Carboxymuconate-epsilon-Sem... -

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Basic information

Entry
Database: PDB / ID: 6mgt
TitleCrystal structure of alpha-Amino-beta-Carboxymuconate-epsilon-Semialdehyde Decarboxylase Mutant H110A
Components2-amino-3-carboxymuconate 6-semialdehyde decarboxylase
KeywordsLYASE / Holo structure / Decarboxylase
Function / homology
Function and homology information


secondary metabolic process / carboxy-lyase activity / hydrolase activity / metal ion binding / cytosol
Similarity search - Function
2-amino-3-carboxymuconate-6-semialdehyde decarboxylase / Amidohydrolase / Amidohydrolase-related / Metal-dependent hydrolases / Metal-dependent hydrolase / TIM Barrel / Alpha-Beta Barrel / Alpha Beta
Similarity search - Domain/homology
: / 2-amino-3-carboxymuconate 6-semialdehyde decarboxylase
Similarity search - Component
Biological speciesPseudomonas fluorescens (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.77 Å
AuthorsYang, Y. / Daivs, I. / Matsui, T. / Rubalcava, I. / Liu, A.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)R01GM108988 United States
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)R21MH10798 United States
National Science Foundation (NSF, United States)CHE-1623856 United States
CitationJournal: J Biol Chem / Year: 2019
Title: Quaternary structure of α-amino-β-carboxymuconate-ϵ-semialdehyde decarboxylase (ACMSD) controls its activity.
Authors: Yu Yang / Ian Davis / Tsutomu Matsui / Ivan Rubalcava / Aimin Liu /
Abstract: α-Amino-β-carboxymuconate-ϵ-semialdehyde decarboxylase (ACMSD) plays an important role in l-tryptophan degradation via the kynurenine pathway. ACMSD forms a homodimer and is functionally inactive ...α-Amino-β-carboxymuconate-ϵ-semialdehyde decarboxylase (ACMSD) plays an important role in l-tryptophan degradation via the kynurenine pathway. ACMSD forms a homodimer and is functionally inactive as a monomer because its catalytic assembly requires an arginine residue from a neighboring subunit. However, how the oligomeric state and self-association of ACMSD are controlled in solution remains unexplored. Here, we demonstrate that ACMSD from can self-assemble into homodimer, tetramer, and higher-order structures. Using size-exclusion chromatography coupled with small-angle X-ray scattering (SEC-SAXS) analysis, we investigated the ACMSD tetramer structure, and fitting the SAXS data with X-ray crystal structures of the monomeric component, we could generate a pseudo-atomic structure of the tetramer. This analysis revealed a tetramer model of ACMSD as a head-on dimer of dimers. We observed that the tetramer is catalytically more active than the dimer and is in equilibrium with the monomer and dimer. Substituting a critical residue of the dimer-dimer interface, His-110, altered the tetramer dissociation profile by increasing the higher-order oligomer portion in solution without changing the X-ray crystal structure. ACMSD self-association was affected by pH, ionic strength, and other electrostatic interactions. Alignment of ACMSD sequences revealed that His-110 is highly conserved in a few bacteria that utilize nitrobenzoic acid as a sole source of carbon and energy, suggesting a dedicated functional role of ACMSD's self-assembly into the tetrameric and higher-order structures. These results indicate that the dynamic oligomerization status potentially regulates ACMSD activity and that SEC-SAXS coupled with X-ray crystallography is a powerful tool for studying protein self-association.
History
DepositionSep 14, 2018Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 19, 2019Provider: repository / Type: Initial release
Revision 1.1Jun 26, 2019Group: Data collection / Database references
Category: citation / citation_author / pdbx_database_related
Item: _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed ..._citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Aug 7, 2019Group: Data collection / Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title / _citation_author.identifier_ORCID
Revision 1.3Nov 20, 2019Group: Derived calculations / Category: pdbx_struct_conn_angle / struct_conn
Revision 1.4Nov 27, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.5Oct 11, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 2-amino-3-carboxymuconate 6-semialdehyde decarboxylase
B: 2-amino-3-carboxymuconate 6-semialdehyde decarboxylase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)79,4184
Polymers79,3012
Non-polymers1182
Water36020
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area5240 Å2
ΔGint-61 kcal/mol
Surface area23500 Å2
MethodPISA
Unit cell
Length a, b, c (Å)90.934, 90.934, 170.107
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number94
Space group name H-MP42212
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11(chain A and (resid 4 through 18 or resid 20...
21(chain B and (resid 4 through 18 or resid 20...

NCS domain segments:
Dom-IDComponent-IDEns-IDSelection detailsAuth asym-IDAuth seq-ID
111(chain A and (resid 4 through 18 or resid 20...A4 - 18
121(chain A and (resid 4 through 18 or resid 20...A20 - 27
131(chain A and (resid 4 through 18 or resid 20...A29 - 36
141(chain A and (resid 4 through 18 or resid 20...A38 - 46
151(chain A and (resid 4 through 18 or resid 20...A55 - 66
161(chain A and (resid 4 through 18 or resid 20...A68 - 91
171(chain A and (resid 4 through 18 or resid 20...A4 - 333
181(chain A and (resid 4 through 18 or resid 20...A4 - 333
191(chain A and (resid 4 through 18 or resid 20...A145 - 158
1101(chain A and (resid 4 through 18 or resid 20...A160 - 164
1111(chain A and (resid 4 through 18 or resid 20...A166 - 1871
1121(chain A and (resid 4 through 18 or resid 20...A303 - 31
1131(chain A and (resid 4 through 18 or resid 20...A303 - 314
1141(chain A and (resid 4 through 18 or resid 20...A316 - 327
1151(chain A and (resid 4 through 18 or resid 20...A329 - 332
211(chain B and (resid 4 through 18 or resid 20...B4 - 18
221(chain B and (resid 4 through 18 or resid 20...B20 - 27
231(chain B and (resid 4 through 18 or resid 20...B29 - 36
241(chain B and (resid 4 through 18 or resid 20...B55 - 66
251(chain B and (resid 4 through 18 or resid 20...B68 - 91
261(chain B and (resid 4 through 18 or resid 20...B68 - 91
271(chain B and (resid 4 through 18 or resid 20...B145 - 158
281(chain B and (resid 4 through 18 or resid 20...B160 - 164
291(chain B and (resid 4 through 18 or resid 20...B166 - 187
2101(chain B and (resid 4 through 18 or resid 20...B189 - 301
2111(chain B and (resid 4 through 18 or resid 20...B303 - 314
2121(chain B and (resid 4 through 18 or resid 20...B316 - 327
2131(chain B and (resid 4 through 18 or resid 20...B329 - 332

NCS oper: (Code: given
Matrix: (-0.99384, 0.074929, -0.081657), (0.071616, -0.128105, -0.989172), (-0.084578, -0.988926, 0.121949)
Vector: 110.647003, 91.601799, 88.975899)

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Components

#1: Protein 2-amino-3-carboxymuconate 6-semialdehyde decarboxylase


Mass: 39650.266 Da / Num. of mol.: 2 / Mutation: H110A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas fluorescens (bacteria) / Gene: nbaD / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q83V25
#2: Chemical ChemComp-CO / COBALT (II) ION


Mass: 58.933 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Co
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 20 / 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: 2.4 Å3/Da / Density % sol: 48.75 %
Crystal growTemperature: 298 K / Method: vapor diffusion, hanging drop / pH: 6
Details: 8% (v/v) TacsimateTM pH 6.0 and 22% (w/v) polyethylene glycol 3,350

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SSRL / Beamline: BL9-2 / Wavelength: 0.97946 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jun 19, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97946 Å / Relative weight: 1
ReflectionResolution: 2.77→50 Å / Num. obs: 17420 / % possible obs: 92 % / Redundancy: 5 % / Biso Wilson estimate: 76.66 Å2 / Rmerge(I) obs: 0.114 / Rpim(I) all: 0.055 / Rrim(I) all: 0.128 / Χ2: 1.019 / Net I/σ(I): 5
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. unique obsCC1/2Rpim(I) allRrim(I) allΧ2% possible all
2.77-2.825.10.9088670.5270.4441.0160.52694.5
2.82-2.875.10.7948670.6070.3890.8890.50394.9
2.87-2.925.10.6938890.6440.3370.7740.54594.6
2.92-2.9850.6198620.70.3020.6920.54493.8
2.98-3.0550.5118850.8060.2480.5710.5594.7
3.05-3.124.90.4358510.8190.2130.4870.58993.1
3.12-3.24.80.3628380.8830.1790.4060.68591
3.2-3.284.40.2698670.9090.1360.3030.71992.3
3.28-3.385.30.2318810.960.1080.2560.893.9
3.38-3.4950.2828560.9410.1460.321.27692.9
3.49-3.615.20.1738690.9720.0810.1921.00793.3
3.61-3.765.10.1678650.9580.0790.1851.29992.7
3.76-3.934.90.1588640.9550.0790.1781.49792.1
3.93-4.145.10.1158820.9870.0540.1281.24492.2
4.14-4.44.90.1048630.9870.0490.1161.37391.8
4.4-4.744.60.0968590.9890.0470.1071.57689.3
4.74-5.2150.098680.990.0420.11.53890.6
5.21-5.965.20.0838780.9910.0380.0921.27490.6
5.96-7.515.10.0758890.9920.0360.0831.25889.6
7.51-504.60.0599200.9950.0290.0671.62784.6

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Processing

Software
NameVersionClassification
DENZOdata reduction
HKL-2000data scaling
PHENIX(1.11.1_2575)refinement
PDB_EXTRACT3.24data extraction
PHENIX1.10.1phasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 2HBV

2hbv


Resolution: 2.77→31.058 Å / SU ML: 0.43 / Cross valid method: THROUGHOUT / σ(F): 1.35 / Phase error: 27.78
RfactorNum. reflection% reflection
Rfree0.2581 1740 10 %
Rwork0.1958 --
obs0.202 17398 92.16 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso max: 145.97 Å2 / Biso mean: 79.3997 Å2 / Biso min: 39.99 Å2
Refinement stepCycle: final / Resolution: 2.77→31.058 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5146 0 2 20 5168
Biso mean--65.54 66.4 -
Num. residues----660
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0095290
X-RAY DIFFRACTIONf_angle_d1.1197169
X-RAY DIFFRACTIONf_chiral_restr0.053765
X-RAY DIFFRACTIONf_plane_restr0.007942
X-RAY DIFFRACTIONf_dihedral_angle_d16.9553153
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDNumberRefine-IDTypeRms dev position (Å)
11A2421X-RAY DIFFRACTIONPOSITIONAL0.063
12B2421X-RAY DIFFRACTIONPOSITIONAL0.063
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 12

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
2.7689-2.85030.36861440.29241305144994
2.8503-2.94220.35291440.26951301144595
2.9422-3.04730.30581460.26751308145494
3.0473-3.16920.32751400.25471264140492
3.1692-3.31330.3531450.251296144193
3.3133-3.48770.30751450.24851308145393
3.4877-3.70590.29491460.21561306145293
3.7059-3.99150.27141430.20151289143292
3.9915-4.39220.25531440.18281302144692
4.3922-5.02540.24111440.16071295143990
5.0254-6.32270.22281470.17881321146891
6.3227-31.06040.19831520.15991363151588

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