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- PDB-6wtx: Structure of VcINDY in complex with terephthalate -

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Basic information

Entry
Database: PDB / ID: 6wtx
TitleStructure of VcINDY in complex with terephthalate
ComponentsDASS family sodium-coupled anion symporter
KeywordsTRANSPORT PROTEIN / Transporter / MEMBRANE PROTEIN
Function / homology
Function and homology information


transmembrane transporter activity / transmembrane transport / identical protein binding / membrane / plasma membrane
Similarity search - Function
Citrate transporter-like domain / Citrate transporter / Sodium:sulfate symporter transmembrane region / Sodium/sulphate symporter, conserved site / Sodium:sulfate symporter family signature. / Solute carrier family 13
Similarity search - Domain/homology
terephthalic acid / Transporter, NadC family / Transporter, NadC family
Similarity search - Component
Biological speciesVibrio cholerae (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.92 Å
AuthorsSauer, D.B. / Marden, J.J. / Wang, D.N.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01NS108151 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM121994 United States
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)R01DK099023 United States
CitationJournal: Elife / Year: 2020
Title: Structural basis for the reaction cycle of DASS dicarboxylate transporters.
Authors: David B Sauer / Noah Trebesch / Jennifer J Marden / Nicolette Cocco / Jinmei Song / Akiko Koide / Shohei Koide / Emad Tajkhorshid / Da-Neng Wang /
Abstract: Citrate, α-ketoglutarate and succinate are TCA cycle intermediates that also play essential roles in metabolic signaling and cellular regulation. These di- and tricarboxylates are imported into the ...Citrate, α-ketoglutarate and succinate are TCA cycle intermediates that also play essential roles in metabolic signaling and cellular regulation. These di- and tricarboxylates are imported into the cell by the divalent anion sodium symporter (DASS) family of plasma membrane transporters, which contains both cotransporters and exchangers. While DASS proteins transport substrates via an elevator mechanism, to date structures are only available for a single DASS cotransporter protein in a substrate-bound, inward-facing state. We report multiple cryo-EM and X-ray structures in four different states, including three hitherto unseen states, along with molecular dynamics simulations, of both a cotransporter and an exchanger. Comparison of these outward- and inward-facing structures reveal how the transport domain translates and rotates within the framework of the scaffold domain through the transport cycle. Additionally, we propose that DASS transporters ensure substrate coupling by a charge-compensation mechanism, and by structural changes upon substrate release.
History
DepositionMay 4, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 16, 2020Provider: repository / Type: Initial release
Revision 1.1Oct 18, 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

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Assembly

Deposited unit
C: DASS family sodium-coupled anion symporter
D: DASS family sodium-coupled anion symporter
A: DASS family sodium-coupled anion symporter
B: DASS family sodium-coupled anion symporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)193,47816
Polymers192,6294
Non-polymers84812
Water00
1
C: DASS family sodium-coupled anion symporter
D: DASS family sodium-coupled anion symporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)96,7398
Polymers96,3152
Non-polymers4246
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
A: DASS family sodium-coupled anion symporter
B: DASS family sodium-coupled anion symporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)96,7398
Polymers96,3152
Non-polymers4246
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)108.458, 103.062, 174.447
Angle α, β, γ (deg.)90.000, 95.848, 90.000
Int Tables number4
Space group name H-MP1211
Space group name HallP2yb
Symmetry operation#1: x,y,z
#2: -x,y+1/2,-z
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "A"
d_2ens_1(chain "B" and resid 18 through 462)
d_3ens_1(chain "C" and resid 18 through 462)
d_4ens_1(chain "D" and resid 18 through 462)

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1LEUGLNE1 - 445
d_21ens_1LEUGLNF1 - 445
d_31ens_1LEUGLNA1 - 445
d_41ens_1LEUGLNC1 - 445

NCS oper:
IDCodeMatrixVector
1given(0.17881747953, -0.26769583762, -0.946764620978), (-0.263848609377, -0.940074780717, 0.215970641501), (-0.947843985234, 0.211183202886, -0.238732977349)-74.3699631073, -48.9424476476, -78.903405897
2given(-0.967565221173, -0.0128845403398, 0.252292551213), (-0.0135317278167, 0.999908097267, -0.000830278188599), (-0.252258667085, -0.00421730243238, -0.967650649377)-24.1344537603, 22.4611615268, -81.8146948528
3given(-0.408632630453, 0.323667314057, 0.853380831247), (-0.262338623631, -0.937204036619, 0.229841337221), (0.874184088096, -0.129954082487, 0.467882588439)28.4875520169, -25.3625225035, 13.5106053603

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Components

#1: Protein
DASS family sodium-coupled anion symporter / Transporter / NadC family


Mass: 48157.359 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae (bacteria) / Gene: VC0395_0108, GG844_00510 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A0H3AG83, UniProt: Q9KNE0*PLUS
#2: Chemical
ChemComp-UB7 / terephthalic acid / benzene-1,4-dicarboxylic acid


Mass: 166.131 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C8H6O4 / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical
ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Na
Has ligand of interestY

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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: 5.04 Å3/Da / Density % sol: 75.57 %
Crystal growTemperature: 277 K / Method: vapor diffusion, hanging drop / pH: 8 / Details: 30% PEG 550, 100mM Tris pH 8.0, 50mM NaCl

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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 PILATUS3 6M / Detector: PIXEL / Date: Nov 2, 2016
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 3.9→50 Å / Num. obs: 33199 / % possible obs: 97.4 % / Redundancy: 4.4 % / Biso Wilson estimate: 7.1 Å2 / Rsym value: 0.112 / Net I/σ(I): 13.3
Reflection shellResolution: 3.9→3.97 Å / Num. unique obs: 1665 / CC1/2: 0.814

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Processing

Software
NameVersionClassification
PHENIX1.18_3855refinement
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5ul9

5ul9


Resolution: 3.92→47.8 Å / SU ML: 0.5189 / Cross valid method: FREE R-VALUE / σ(F): 1.36 / Phase error: 30.1207
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.3084 2000 6.98 %
Rwork0.2902 26638 -
obs0.2915 28638 82.4 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 5.64 Å2
Refinement stepCycle: LAST / Resolution: 3.92→47.8 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms13372 0 56 0 13428
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.005913736
X-RAY DIFFRACTIONf_angle_d1.16618728
X-RAY DIFFRACTIONf_chiral_restr0.06652320
X-RAY DIFFRACTIONf_plane_restr0.01032268
X-RAY DIFFRACTIONf_dihedral_angle_d6.07351824
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefine-IDTypeRms dev position (Å)
ens_1d_2BX-RAY DIFFRACTIONTorsion NCS0.347817121648
ens_1d_3CX-RAY DIFFRACTIONTorsion NCS0.251664234019
ens_1d_4DX-RAY DIFFRACTIONTorsion NCS0.30342712955
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.92-4.010.4453820.3531082X-RAY DIFFRACTION46.97
4.01-4.120.33071030.32571390X-RAY DIFFRACTION60.99
4.12-4.240.35751140.31251517X-RAY DIFFRACTION66.41
4.24-4.380.33341240.29931660X-RAY DIFFRACTION72.64
4.38-4.540.29011350.29181788X-RAY DIFFRACTION77.76
4.54-4.720.30331430.28671913X-RAY DIFFRACTION82.64
4.72-4.930.3011510.27592006X-RAY DIFFRACTION87.79
4.93-5.190.30581570.29292099X-RAY DIFFRACTION91.04
5.19-5.520.30341670.30082225X-RAY DIFFRACTION97
5.52-5.940.31511730.31192295X-RAY DIFFRACTION99.52
5.94-6.540.34751720.28542300X-RAY DIFFRACTION99.56
6.54-7.480.2441730.26332303X-RAY DIFFRACTION98.92
7.49-9.420.22541590.21322118X-RAY DIFFRACTION90.75
9.42-47.80.29891470.29231942X-RAY DIFFRACTION81.28

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