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- PDB-4cdi: Crystal structure of AcrB-AcrZ complex -

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Basic information

Entry
Database: PDB / ID: 4cdi
TitleCrystal structure of AcrB-AcrZ complex
Components
  • ACRIFLAVINE RESISTANCE PROTEIN B
  • PREDICTED PROTEIN
KeywordsMEMBRANE PROTEIN / DRUG EFFLUX / TRANSMEMBRANE PROTEIN
Function / homology
Function and homology information


xenobiotic detoxification by transmembrane export across the cell outer membrane / efflux pump complex / periplasmic side of plasma membrane / efflux transmembrane transporter activity / xenobiotic transmembrane transporter activity / outer membrane-bounded periplasmic space / identical protein binding / membrane / plasma membrane
Similarity search - Function
Single alpha-helices involved in coiled-coils or other helix-helix interfaces - #2480 / Multidrug efflux transporter AcrB transmembrane fold / Multidrug efflux transporter AcrB transmembrane domain / Multidrug efflux transporter AcrB TolC docking domain; DN and DC subdomains / Multidrug efflux transporter AcrB TolC docking domain; DN and DC subdomains / Multidrug efflux transporter AcrB pore domain / Multidrug efflux transporter AcrB pore domain like / Multidrug efflux transporter AcrB pore domain / Hydrophobe/amphiphile efflux-1 HAE1 / Acriflavin resistance protein ...Single alpha-helices involved in coiled-coils or other helix-helix interfaces - #2480 / Multidrug efflux transporter AcrB transmembrane fold / Multidrug efflux transporter AcrB transmembrane domain / Multidrug efflux transporter AcrB TolC docking domain; DN and DC subdomains / Multidrug efflux transporter AcrB TolC docking domain; DN and DC subdomains / Multidrug efflux transporter AcrB pore domain / Multidrug efflux transporter AcrB pore domain like / Multidrug efflux transporter AcrB pore domain / Hydrophobe/amphiphile efflux-1 HAE1 / Acriflavin resistance protein / Multidrug efflux transporter AcrB TolC docking domain, DN/DC subdomains / AcrB/AcrD/AcrF family / Single alpha-helices involved in coiled-coils or other helix-helix interfaces / Helix non-globular / Special / Alpha-Beta Plaits / Up-down Bundle / 2-Layer Sandwich / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
: / Multidrug efflux pump subunit AcrB
Similarity search - Component
Biological speciesESCHERICHIA COLI (E. coli)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.7 Å
AuthorsDu, D. / James, N. / Klimont, E. / Luisi, B.F.
CitationJournal: Nature / Year: 2014
Title: Structure of the AcrAB-TolC multidrug efflux pump.
Authors: Dijun Du / Zhao Wang / Nathan R James / Jarrod E Voss / Ewa Klimont / Thelma Ohene-Agyei / Henrietta Venter / Wah Chiu / Ben F Luisi /
Abstract: The capacity of numerous bacterial species to tolerate antibiotics and other toxic compounds arises in part from the activity of energy-dependent transporters. In Gram-negative bacteria, many of ...The capacity of numerous bacterial species to tolerate antibiotics and other toxic compounds arises in part from the activity of energy-dependent transporters. In Gram-negative bacteria, many of these transporters form multicomponent 'pumps' that span both inner and outer membranes and are driven energetically by a primary or secondary transporter component. A model system for such a pump is the acridine resistance complex of Escherichia coli. This pump assembly comprises the outer-membrane channel TolC, the secondary transporter AcrB located in the inner membrane, and the periplasmic AcrA, which bridges these two integral membrane proteins. The AcrAB-TolC efflux pump is able to transport vectorially a diverse array of compounds with little chemical similarity, thus conferring resistance to a broad spectrum of antibiotics. Homologous complexes are found in many Gram-negative species, including in animal and plant pathogens. Crystal structures are available for the individual components of the pump and have provided insights into substrate recognition, energy coupling and the transduction of conformational changes associated with the transport process. However, how the subunits are organized in the pump, their stoichiometry and the details of their interactions are not known. Here we present the pseudo-atomic structure of a complete multidrug efflux pump in complex with a modulatory protein partner from E. coli. The model defines the quaternary organization of the pump, identifies key domain interactions, and suggests a cooperative process for channel assembly and opening. These findings illuminate the basis for drug resistance in numerous pathogenic bacterial species.
History
DepositionOct 31, 2013Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 30, 2014Provider: repository / Type: Initial release
Revision 1.1May 28, 2014Group: Database references
Revision 1.2Sep 23, 2015Group: Data collection
Revision 1.3May 8, 2019Group: Data collection / Experimental preparation / Other
Category: exptl_crystal_grow / pdbx_database_proc / pdbx_database_status
Item: _exptl_crystal_grow.temp / _pdbx_database_status.recvd_author_approval
Revision 1.4May 29, 2019Group: Data collection / Experimental preparation / Category: exptl_crystal_grow / struct_biol / Item: _exptl_crystal_grow.method
Revision 1.5Dec 20, 2023Group: Data collection / Database references ...Data collection / Database references / Other / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_database_status / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_database_status.status_code_sf

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: ACRIFLAVINE RESISTANCE PROTEIN B
C: PREDICTED PROTEIN


Theoretical massNumber of molelcules
Total (without water)118,9702
Polymers118,9702
Non-polymers00
Water00
1
A: ACRIFLAVINE RESISTANCE PROTEIN B
C: PREDICTED PROTEIN

A: ACRIFLAVINE RESISTANCE PROTEIN B
C: PREDICTED PROTEIN

A: ACRIFLAVINE RESISTANCE PROTEIN B
C: PREDICTED PROTEIN


Theoretical massNumber of molelcules
Total (without water)356,9096
Polymers356,9096
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_445-y-1,x-y-1,z1
crystal symmetry operation3_545-x+y,-x-1,z1
Buried area27120 Å2
ΔGint-167.5 kcal/mol
Surface area121710 Å2
MethodPISA
Unit cell
Length a, b, c (Å)146.205, 146.205, 543.207
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number155
Space group name H-MH32

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Components

#1: Protein ACRIFLAVINE RESISTANCE PROTEIN B / ACRB


Mass: 113665.180 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) ESCHERICHIA COLI (E. coli) / Description: NOVAGENE / Variant: NOVABLUE ISOLATE / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): BL21(DE3) / Variant (production host): C43 / References: UniProt: P31224
#2: Protein/peptide PREDICTED PROTEIN / ACRZ


Mass: 5304.423 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) ESCHERICHIA COLI (E. coli) / Strain: W3110 / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): BL21(DE3) / Variant (production host): C43 / References: UniProt: C4ZXT3

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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: 4.51 Å3/Da / Density % sol: 72.8 % / Description: NONE
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop / pH: 7.4
Details: THE ACRBZ COMPLEX AT 10 MG ML-1 USING SAMPLE BUFFER. 9 MM N-OCTYL-BETA-D-THIOGLUCOPYRANOSIDE (90 MM) WAS MIXED WITH ACRBZ COMPLEX BEFORE THE CRYSTALLISATION TRIALS. THE ACRBZ CRYSTALS WERE ...Details: THE ACRBZ COMPLEX AT 10 MG ML-1 USING SAMPLE BUFFER. 9 MM N-OCTYL-BETA-D-THIOGLUCOPYRANOSIDE (90 MM) WAS MIXED WITH ACRBZ COMPLEX BEFORE THE CRYSTALLISATION TRIALS. THE ACRBZ CRYSTALS WERE GROWN AT 20 C USING THE HANGING-DROPLET VAPOUR DIFFUSION METHOD BY MIXING 4 MICROLITERS OF ACRBZ COMPLEX WITH 2 MICROLITERS OF RESERVOIR SOLUTION (100 MM TRICINE PH: 7.4, 50 MM LITHIUM SULPHATE, 5 MM CADMIUM CHLORIDE HYDRATE, 7 % PEG 3000, 10% GLYCEROL).

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Data collection

Diffraction
IDMean temperature (K)Crystal-ID
11001
21001
Diffraction source
SourceSiteBeamlineIDWavelength
SYNCHROTRONDiamond I2410.9794
SYNCHROTRONDiamond I04-120.9794
Detector
TypeIDDetectorDate
DECTRIS PILATUS 2M1PIXELJan 10, 2013
DECTRIS PIXEL2PIXELJan 10, 2013
Radiation
IDProtocolMonochromatic (M) / Laue (L)Scattering typeWavelength-ID
1SINGLE WAVELENGTHMx-ray1
2SINGLE WAVELENGTHMx-ray1
Radiation wavelengthWavelength: 0.9794 Å / Relative weight: 1
ReflectionResolution: 3.69→24.89 Å / Num. obs: 23077 / % possible obs: 94.5 % / Observed criterion σ(I): 2 / Redundancy: 10.6 % / Biso Wilson estimate: 148 Å2 / Rmerge(I) obs: 0.13 / Net I/σ(I): 15.1
Reflection shellResolution: 3.69→3.99 Å / Redundancy: 10.6 % / Rmerge(I) obs: 1.17 / Mean I/σ(I) obs: 2.1 / % possible all: 94.8

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Processing

Software
NameVersionClassification
REFMAC5.7.0032refinement
MOSFLMdata reduction
SCALAdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 4C48

4c48


Resolution: 3.7→24.91 Å / Cor.coef. Fo:Fc: 0.864 / Cor.coef. Fo:Fc free: 0.858 / SU B: 54.668 / SU ML: 0.78 / Cross valid method: THROUGHOUT / ESU R Free: 0.844 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS.
RfactorNum. reflection% reflectionSelection details
Rfree0.36127 1161 5 %RANDOM
Rwork0.33906 ---
obs0.34009 21912 94.04 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 117.759 Å2
Baniso -1Baniso -2Baniso -3
1-3.41 Å23.41 Å20 Å2
2--3.41 Å20 Å2
3----11.07 Å2
Refinement stepCycle: LAST / Resolution: 3.7→24.91 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms8114 0 0 0 8114
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.010.0198266
X-RAY DIFFRACTIONr_bond_other_d0.0020.028103
X-RAY DIFFRACTIONr_angle_refined_deg1.3981.9711228
X-RAY DIFFRACTIONr_angle_other_deg0.818318582
X-RAY DIFFRACTIONr_dihedral_angle_1_deg7.8851075
X-RAY DIFFRACTIONr_dihedral_angle_2_deg35.18824.583312
X-RAY DIFFRACTIONr_dihedral_angle_3_deg18.157151374
X-RAY DIFFRACTIONr_dihedral_angle_4_deg12.2951535
X-RAY DIFFRACTIONr_chiral_restr0.0720.21330
X-RAY DIFFRACTIONr_gen_planes_refined0.0060.0219370
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021807
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_it1.12311.9684309
X-RAY DIFFRACTIONr_mcbond_other1.12311.9684308
X-RAY DIFFRACTIONr_mcangle_it1.58217.9555381
X-RAY DIFFRACTIONr_mcangle_other
X-RAY DIFFRACTIONr_scbond_it1.11411.9453956
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: 3.695→3.789 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.392 86 -
Rwork0.42 1544 -
obs--93.62 %

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