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Basic information

Entry
Database: PDB / ID: 4afd
TitleStructural and biochemical characterization of a novel Carbohydrate Binding Module of endoglucanase Cel5A from Eubacterium cellulosolvens with a partially bound cellotetraose moeity.
ComponentsENDOGLUCANASE CEL5A
KeywordsHYDROLASE / FAMILY 5 GLYCOSIDE HYDROLASE / CELLULOSOME / CELLOTETRAOSE
Function / homology
Function and homology information


glucan catabolic process / beta-glucosidase activity / cell surface / extracellular region
Similarity search - Function
Jelly Rolls - #1070 / Carbohydrate binding module 65 domain 1 / Carbohydrate binding module 65 domain 1 / Glycoside hydrolase, family 5 / Cellulase (glycosyl hydrolase family 5) / Glycoside hydrolase superfamily / Jelly Rolls / Sandwich / Mainly Beta
Similarity search - Domain/homology
beta-cellotetraose / Endoglucanase cel5A
Similarity search - Component
Biological speciesEUBACTERIUM CELLULOSOLVENS (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.34 Å
AuthorsLuis, A.S. / Venditto, I. / Prates, J.A.M. / Ferreira, L.M.A. / Gilbert, H.J. / Fontes, C.M.G.A. / Najmudin, S.
Citation
Journal: J.Biol.Chem. / Year: 2013
Title: Understanding How Noncatalytic Carbohydrate Binding Modules Can Display Specificity for Xyloglucan.
Authors: Luis, A.S. / Venditto, I. / Temple, M.J. / Rogowski, A. / Basle, A. / Xue, J. / Knox, J.P. / Prates, J.A.M. / Ferreira, L.M.A. / Fontes, C.M.G.A. / Najmudin, S. / Gilbert, H.J.
#1: Journal: Acta Crystallogr.,Sect.F / Year: 2011
Title: Overproduction, Purification, Crystallization and Preliminary X-Ray Characterization of a Novel Carbohydrate-Binding Module of Endoglucanase Cel5A from Eubacterium Cellulosolvens.
Authors: Luis, A.S. / Alves, V.D. / Romao, M.J. / Prates, J.A.M. / Fontes, C.M.G.A. / Najmudin, S.
History
DepositionJan 18, 2012Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 26, 2012Provider: repository / Type: Initial release
Revision 1.1Feb 19, 2014Group: Database references
Revision 1.2May 8, 2019Group: Data collection / Derived calculations ...Data collection / Derived calculations / Experimental preparation / Other
Category: exptl_crystal_grow / pdbx_database_proc ...exptl_crystal_grow / pdbx_database_proc / pdbx_database_status / pdbx_struct_special_symmetry / struct_conn
Item: _exptl_crystal_grow.temp / _pdbx_database_status.recvd_author_approval / _struct_conn.pdbx_leaving_atom_flag
Revision 2.0Jul 29, 2020Group: Advisory / Atomic model ...Advisory / Atomic model / Data collection / Derived calculations / Other / Structure summary
Category: atom_site / chem_comp ...atom_site / chem_comp / database_PDB_caveat / entity / entity_name_com / pdbx_branch_scheme / pdbx_chem_comp_identifier / pdbx_database_status / pdbx_entity_branch / pdbx_entity_branch_descriptor / pdbx_entity_branch_link / pdbx_entity_branch_list / pdbx_entity_nonpoly / pdbx_molecule_features / pdbx_nonpoly_scheme / pdbx_struct_assembly_gen / pdbx_struct_special_symmetry / pdbx_validate_chiral / struct_asym / struct_conn / struct_site / struct_site_gen
Item: _atom_site.auth_asym_id / _atom_site.auth_seq_id ..._atom_site.auth_asym_id / _atom_site.auth_seq_id / _atom_site.label_asym_id / _chem_comp.name / _chem_comp.type / _entity.formula_weight / _entity.pdbx_description / _entity.pdbx_number_of_molecules / _entity.type / _pdbx_database_status.status_code_sf / _pdbx_struct_assembly_gen.asym_id_list / _pdbx_struct_special_symmetry.label_asym_id / _pdbx_validate_chiral.auth_asym_id / _pdbx_validate_chiral.auth_seq_id / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id
Description: Carbohydrate remediation / Provider: repository / Type: Remediation
Revision 2.1Dec 20, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description / Structure summary
Category: chem_comp / chem_comp_atom ...chem_comp / chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_conn
Item: _chem_comp.pdbx_synonyms / _database_2.pdbx_DOI ..._chem_comp.pdbx_synonyms / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.pdbx_leaving_atom_flag

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: ENDOGLUCANASE CEL5A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)15,5232
Polymers14,8561
Non-polymers6671
Water3,639202
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
Unit cell
Length a, b, c (Å)48.773, 48.773, 193.719
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number178
Space group name H-MP6122
Components on special symmetry positions
IDModelComponents
11A-2036-

HOH

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Components

#1: Protein ENDOGLUCANASE CEL5A


Mass: 14856.365 Da / Num. of mol.: 1
Fragment: N-TERMINAL CARBOHYDRATE BINDING MODULE, RESIDUES 40-170
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) EUBACTERIUM CELLULOSOLVENS (bacteria) / Production host: ESCHERICHIA COLI (E. coli) / References: UniProt: Q3LHN3
#2: Polysaccharide beta-D-glucopyranose-(1-4)-beta-D-glucopyranose-(1-4)-beta-D-glucopyranose-(1-4)-beta-D-glucopyranose / beta-cellotetraose


Type: oligosaccharide, Oligosaccharide / Class: Metabolism / Mass: 666.578 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: oligosaccharide / References: beta-cellotetraose
DescriptorTypeProgram
DGlcpb1-4DGlcpb1-4DGlcpb1-4DGlcpb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,4,3/[a2122h-1b_1-5]/1-1-1-1/a4-b1_b4-c1_c4-d1WURCSPDB2Glycan 1.1.0
[][b-D-Glcp]{[(4+1)][b-D-Glcp]{[(4+1)][b-D-Glcp]{[(4+1)][b-D-Glcp]{}}}}LINUCSPDB-CARE
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 202 / Source method: isolated from a natural source / Formula: H2O
Nonpolymer detailsBETA-D-GLUCOSE (BGC): THE LIGAND IS CELLOTETRAOSE MOIETY OF A CELLOHEXAOSE SUBSTRATE USED IN A ...BETA-D-GLUCOSE (BGC): THE LIGAND IS CELLOTETRAOSE MOIETY OF A CELLOHEXAOSE SUBSTRATE USED IN A COCRYSTALLISATION EXPERIMENT, IE B-D-GLUCOPYRANOSYL-1,4-B-D-GLUCOPYRANOSYL-1, 4-B-D-GLUCOPYRANOSYL-1,4-B-D-GLUCOPYRANOSE.

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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: 2.23 Å3/Da / Density % sol: 45.05 % / Description: NONE
Crystal growTemperature: 292 K / pH: 4.6
Details: 101 MG/ML PROTEIN COCRYSTALLISED WITH 1 0 MM CELLOHEXAOSE AT 292 K WITH 0.2 M AMMONIUM SULFATE, 0.1 M SODIUM ACETATE TRIHYDRATE PH 4.6, 26% W/V PEG 2K MME. 30% GLYCEROL WAS USED AS ...Details: 101 MG/ML PROTEIN COCRYSTALLISED WITH 1 0 MM CELLOHEXAOSE AT 292 K WITH 0.2 M AMMONIUM SULFATE, 0.1 M SODIUM ACETATE TRIHYDRATE PH 4.6, 26% W/V PEG 2K MME. 30% GLYCEROL WAS USED AS CRYOPROTECTANT IN MOTHER LIQUOR SOLUTION.

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SOLEIL / Beamline: PROXIMA 1 / Wavelength: 0.98011
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Oct 7, 2011
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.98011 Å / Relative weight: 1
ReflectionResolution: 1.34→42.24 Å / Num. obs: 30670 / % possible obs: 96.9 % / Redundancy: 10.7 % / Rmerge(I) obs: 0.05 / Net I/σ(I): 26.3
Reflection shellResolution: 1.34→1.41 Å / Redundancy: 10.8 % / Rmerge(I) obs: 0.48 / Mean I/σ(I) obs: 5 / % possible all: 93.6

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Processing

Software
NameVersionClassification
REFMAC5.6.0117refinement
XDSdata reduction
SCALAdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 4AEK

4aek


Resolution: 1.34→42.24 Å / Cor.coef. Fo:Fc: 0.965 / Cor.coef. Fo:Fc free: 0.96 / SU B: 1.394 / SU ML: 0.029 / Cross valid method: THROUGHOUT / ESU R: 0.054 / ESU R Free: 0.055 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS. HYDROGENS HAVE BEEN USED IF PRESENT IN THE INPUT. U VALUES REFINED INDIVIDUALLY. ASP A118 HAS BEEN GIVEN DUAL CONFORMATION. IN ONE FORM IT ...Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS. HYDROGENS HAVE BEEN USED IF PRESENT IN THE INPUT. U VALUES REFINED INDIVIDUALLY. ASP A118 HAS BEEN GIVEN DUAL CONFORMATION. IN ONE FORM IT APPEARS TO HAVE REACTED WITH THE NH OF GLY A119 TO FORM A CYCLIC ENTITY, POSSIBLY OWING TO RADIATION DAMAGE. SER A45, GLU A51, ASP A59, GLN A67, GLN A71, ILE A83, GLU A100, LYS A104, ILE A111, SER A112, VAL A116, LYS A130, VAL A160, AND SER A112 ALSO HAVE ALTERNATE CONFORMATIONS. PHENIX-1.7.2-869 WAS ALSO USED FOR OCCUPANCY REFINEMENT AND FOR TLS GROUP DETERMINATION AS A PENULTIMATE STEP. RESIDUES A38 AND A166-170 ARE DISORDERED. DISORDERED SIDE CHAINS WERE MODELED STEREOCHEMICALLY. THIS DATASET HAS SPURIOUS ELECTRON DENSITY ACROSS THE POTENTIAL BINDING CLEFT. INITIALLY, A SINGLE GLUCOSE SUBUNIT WAS MODELLED INTO THE DENSITY BETWEEN TRP 108 AND THE SYMMETRY-RELATED TYR 114 AND THE MODEL WAS REFINED. SUBSEQUENTLY, A SECOND GLUCOSE SUBUNIT WAS MODELLED IN THE ELECTRON DENSITY ABOVE TRP 55. BASED ON THESE TWO SUBUNITS, A CELLOTETRAOSE MOIETY WAS DOCKED ON TO THE SURFACE AND THE LIGAND FITTING PROGRAM IN ARP/WARP WAS USED TO FIT IT BASED ON THE ELECTRON DENSITY. THIS IT DID WITH A MAP DENSITY CORRELATION OF 0.73. THE CELLOTETRAOSE MOIETY (BGC A1167-A1170) CAN ALSO BE BUILT EQUALLY WELL IN THE OPPOSITE ORIENTATION AND SHOULD BE REGARDED AS WEAKLY/PARTIALLY BOUND.
RfactorNum. reflection% reflectionSelection details
Rfree0.18681 1522 5 %RANDOM
Rwork0.16548 ---
obs0.16655 29148 96.11 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 15.58 Å2
Baniso -1Baniso -2Baniso -3
1-0.05 Å20.02 Å20 Å2
2--0.05 Å20 Å2
3----0.07 Å2
Refinement stepCycle: LAST / Resolution: 1.34→42.24 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1009 0 45 202 1256
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0290.021177
X-RAY DIFFRACTIONr_bond_other_d
X-RAY DIFFRACTIONr_angle_refined_deg2.6231.971627
X-RAY DIFFRACTIONr_angle_other_deg
X-RAY DIFFRACTIONr_dihedral_angle_1_deg8.6945155
X-RAY DIFFRACTIONr_dihedral_angle_2_deg40.18525.57752
X-RAY DIFFRACTIONr_dihedral_angle_3_deg12.48515182
X-RAY DIFFRACTIONr_dihedral_angle_4_deg11.176151
X-RAY DIFFRACTIONr_chiral_restr0.1560.2187
X-RAY DIFFRACTIONr_gen_planes_refined0.0160.021894
X-RAY DIFFRACTIONr_gen_planes_other
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it
X-RAY DIFFRACTIONr_mcbond_other
X-RAY DIFFRACTIONr_mcangle_it
X-RAY DIFFRACTIONr_mcangle_other
X-RAY DIFFRACTIONr_scbond_it
X-RAY DIFFRACTIONr_scbond_other
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other
X-RAY DIFFRACTIONr_long_range_B_refined
X-RAY DIFFRACTIONr_long_range_B_other
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.34→1.374 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.271 78 -
Rwork0.249 1790 -
obs--91.17 %
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)
10.529-0.6278-0.5310.93331.121.814-0.03130.00430.00520.0080.00370.0069-0.06150.01440.02760.0455-0.0063-0.00150.00530.00350.0365-26.07238.158-3.2357
20.7110.35951.04760.80630.8811.7417-0.02140.1568-0.02570.05890.0817-0.04780.00610.2324-0.06030.0247-0.0107-0.00220.0499-0.0070.0351-13.243830.6699-11.206
30.33480.00310.12030.03010.06530.3545-0.0017-0.024-0.0054-0.00150.0166-0.00930.0106-0.0237-0.01490.046-0.0002-0.00540.0252-0.00050.0362-18.594932.51533.1607
41.7406-0.34050.71420.5330.11320.48640.0370.0216-0.05430.02540.0208-0.05150.0310.0427-0.05780.0270.0038-0.00840.01010.00150.0536-4.988826.46022.7854
50.53330.03360.12490.0789-0.03110.1683-0.0586-0.0327-0.017300.03680.01440.021-0.00840.02180.03990.0105-0.00240.02590.00870.0437-18.431826.20764.0433
60.3505-0.226-0.17210.31380.54751.2249-0.0418-0.0073-0.030.0520.01140.03060.09480.01730.03040.0518-0.00990.00080.01610.00560.0384-24.946331.162-4.0811
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A38 - 50
2X-RAY DIFFRACTION2A51 - 66
3X-RAY DIFFRACTION3A67 - 98
4X-RAY DIFFRACTION4A99 - 112
5X-RAY DIFFRACTION5A113 - 147
6X-RAY DIFFRACTION6A148 - 165

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