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- PDB-6zaz: Fructo-oligosaccharide transporter BT 1762-63 -

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Basic information

Entry
Database: PDB / ID: 6zaz
TitleFructo-oligosaccharide transporter BT 1762-63
Components
  • SusC homolog
  • SusD homolog
KeywordsMEMBRANE PROTEIN / Outer membrane protein TonB-dependent transporter SusC SusD Levan Bacteroides thetaiotaomicron
Function / homology
Function and homology information


cell outer membrane / membrane / metal ion binding
Similarity search - Function
TonB-dependent outer membrane protein, SusC/RagA / TonB-dependent outer membrane protein SusC/RagA, conserved site / CarboxypepD_reg-like domain / SusD-like, N-terminal / Starch-binding associating with outer membrane / RagB/SusD domain / SusD family / Carboxypeptidase-like, regulatory domain superfamily / TonB-dependent receptor-like, beta-barrel / TonB dependent receptor ...TonB-dependent outer membrane protein, SusC/RagA / TonB-dependent outer membrane protein SusC/RagA, conserved site / CarboxypepD_reg-like domain / SusD-like, N-terminal / Starch-binding associating with outer membrane / RagB/SusD domain / SusD family / Carboxypeptidase-like, regulatory domain superfamily / TonB-dependent receptor-like, beta-barrel / TonB dependent receptor / TonB-dependent receptor, plug domain superfamily / TonB-dependent receptor, plug domain / TonB-dependent receptor-like, beta-barrel domain superfamily / TonB-dependent Receptor Plug Domain / Prokaryotic membrane lipoprotein lipid attachment site profile. / Tetratricopeptide-like helical domain superfamily
Similarity search - Domain/homology
SusC homolog / SusD homolog
Similarity search - Component
Biological speciesBacteroides thetaiotaomicron (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.69 Å
Authorsvan den Berg, B.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/P003192/1 United Kingdom
CitationJournal: Nat Commun / Year: 2021
Title: Insights into SusCD-mediated glycan import by a prominent gut symbiont.
Authors: Declan A Gray / Joshua B R White / Abraham O Oluwole / Parthasarathi Rath / Amy J Glenwright / Adam Mazur / Michael Zahn / Arnaud Baslé / Carl Morland / Sasha L Evans / Alan Cartmell / ...Authors: Declan A Gray / Joshua B R White / Abraham O Oluwole / Parthasarathi Rath / Amy J Glenwright / Adam Mazur / Michael Zahn / Arnaud Baslé / Carl Morland / Sasha L Evans / Alan Cartmell / Carol V Robinson / Sebastian Hiller / Neil A Ranson / David N Bolam / Bert van den Berg /
Abstract: In Bacteroidetes, one of the dominant phyla of the mammalian gut, active uptake of large nutrients across the outer membrane is mediated by SusCD protein complexes via a "pedal bin" transport ...In Bacteroidetes, one of the dominant phyla of the mammalian gut, active uptake of large nutrients across the outer membrane is mediated by SusCD protein complexes via a "pedal bin" transport mechanism. However, many features of SusCD function in glycan uptake remain unclear, including ligand binding, the role of the SusD lid and the size limit for substrate transport. Here we characterise the β2,6 fructo-oligosaccharide (FOS) importing SusCD from Bacteroides thetaiotaomicron (Bt1762-Bt1763) to shed light on SusCD function. Co-crystal structures reveal residues involved in glycan recognition and suggest that the large binding cavity can accommodate several substrate molecules, each up to ~2.5 kDa in size, a finding supported by native mass spectrometry and isothermal titration calorimetry. Mutational studies in vivo provide functional insights into the key structural features of the SusCD apparatus and cryo-EM of the intact dimeric SusCD complex reveals several distinct states of the transporter, directly visualising the dynamics of the pedal bin transport mechanism.
History
DepositionJun 6, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 18, 2020Provider: repository / Type: Initial release
Revision 1.1Jan 20, 2021Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _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
Revision 1.2Jan 24, 2024Group: 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: SusD homolog
B: SusC homolog
hetero molecules


Theoretical massNumber of molelcules
Total (without water)184,44910
Polymers181,6262
Non-polymers2,8238
Water4,378243
1
A: SusD homolog
B: SusC homolog
hetero molecules

A: SusD homolog
B: SusC homolog
hetero molecules


Theoretical massNumber of molelcules
Total (without water)368,89820
Polymers363,2524
Non-polymers5,64616
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation3_655-x+1,y,-z+1/21
Buried area35150 Å2
ΔGint-25 kcal/mol
Surface area100510 Å2
MethodPISA
Unit cell
Length a, b, c (Å)119.566, 237.835, 170.505
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number20
Space group name H-MC2221

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Components

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Protein , 2 types, 2 molecules AB

#1: Protein SusD homolog


Mass: 66142.625 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Bacteroides thetaiotaomicron (strain ATCC 29148 / DSM 2079 / NCTC 10582 / E50 / VPI-5482) (bacteria)
Production host: Bacteroides thetaiotaomicron (bacteria) / References: UniProt: Q8A6W4
#2: Protein SusC homolog


Mass: 115483.602 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Bacteroides thetaiotaomicron (strain ATCC 29148 / DSM 2079 / NCTC 10582 / E50 / VPI-5482) (bacteria)
References: UniProt: Q8A6W3

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Sugars , 2 types, 2 molecules

#3: Polysaccharide beta-D-fructofuranose-(2-6)-beta-D-fructofuranose-(2-6)-beta-D-fructofuranose-(2-6)-[beta-D- ...beta-D-fructofuranose-(2-6)-beta-D-fructofuranose-(2-6)-beta-D-fructofuranose-(2-6)-[beta-D-fructofuranose-(2-1)]beta-D-fructofuranose-(2-6)-beta-D-fructofuranose-(2-6)-beta-D-fructofuranose


Type: oligosaccharide / Mass: 1153.001 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DFrufb2-6DFrufb2-6DFrufb2-6[DFrufb2-1]DFrufb2-6DFrufb2-6DFrufb2-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/1,7,6/[ha122h-2b_2-5]/1-1-1-1-1-1-1/a6-b2_b6-c2_c1-d2_c6-e2_e6-f2_f6-g2WURCSPDB2Glycan 1.1.0
[][b-D-Fruf]{[(6+2)][b-D-Fruf]{[(6+2)][b-D-Fruf]{[(6+2)][b-D-Fruf]{[(6+2)][b-D-Fruf]{[(6+2)][b-D-Fruf]{}}}}}}LINUCSPDB-CARE
#4: Polysaccharide beta-D-fructofuranose-(2-1)-beta-D-fructofuranose-(2-6)-beta-D-fructofuranose-(2-6)-beta-D-fructofuranose


Type: oligosaccharide / Mass: 666.578 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DFrufb2-1DFrufb2-6DFrufb2-6DFrufb2-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/1,4,3/[ha122h-2b_2-5]/1-1-1-1/a6-b2_b6-c2_c1-d2WURCSPDB2Glycan 1.1.0
[][b-D-Fruf]{[(6+2)][b-D-Fruf]{[(6+2)][b-D-Fruf]{}}}LINUCSPDB-CARE

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Non-polymers , 4 types, 249 molecules

#5: Chemical ChemComp-CL / CHLORIDE ION / Chloride


Mass: 35.453 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Cl
#6: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#7: Chemical ChemComp-C8E / (HYDROXYETHYLOXY)TRI(ETHYLOXY)OCTANE


Mass: 306.438 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Formula: C16H34O5 / Comment: C8E, detergent*YM
#8: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 243 / Source method: isolated from a natural source / Formula: H2O

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Details

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: 3.65 Å3/Da / Density % sol: 66.33 %
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop
Details: 0.15 M sodium formate 0.1 M Hepes pH 7-7.5 16-20% PEG3350

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: Diamond / Beamline: I04 / Wavelength: 0.978 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Jun 25, 2016
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.978 Å / Relative weight: 1
ReflectionResolution: 2.69→170.5 Å / Num. obs: 67688 / % possible obs: 100 % / Redundancy: 7.2 % / CC1/2: 0.99 / Rpim(I) all: 0.085 / Net I/σ(I): 6
Reflection shellResolution: 2.69→2.74 Å / Mean I/σ(I) obs: 1.1 / Num. unique obs: 3382 / CC1/2: 0.6 / Rpim(I) all: 0.61

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Processing

Software
NameVersionClassification
PHENIX(1.11.1_2575: ???)refinement
xia2data reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5T3R

5t3r


Resolution: 2.69→106.826 Å / SU ML: 0.49 / Cross valid method: FREE R-VALUE / σ(F): 1.33 / Phase error: 32.47 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2753 3304 4.93 %
Rwork0.2083 --
obs0.2117 66963 98.97 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.69→106.826 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms11700 0 175 243 12118
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00812159
X-RAY DIFFRACTIONf_angle_d1.10516501
X-RAY DIFFRACTIONf_dihedral_angle_d7.0537149
X-RAY DIFFRACTIONf_chiral_restr0.0571738
X-RAY DIFFRACTIONf_plane_restr0.0062140
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
10.237-0.00280.16780.2806-0.00020.27710.0996-0.1394-0.10390.0534-0.0113-0.01150.1059-0.00670.00320.2657-0.0485-0.05880.2350.10840.263571.236926.497968.4359
20.1666-0.09760.10060.1488-0.03210.29380.0562-0.06250.03290.028-0.0366-0.0321-0.10560.092600.2168-0.0710.0360.2223-0.03690.223677.509272.449262.5654
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1(chain 'A' and resid 1 through 552)
2X-RAY DIFFRACTION2(chain 'B' and resid 96 through 1016)

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