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- EMDB-4774: Heterodimeric ABC exporter TmrAB in inward-facing wide conformati... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-4774 | ||||||||||||
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Title | Heterodimeric ABC exporter TmrAB in inward-facing wide conformation under turnover conditions | ||||||||||||
![]() | TmrAB in IF wide conformation under turnover conditions | ||||||||||||
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Function / homology | ![]() ATPase-coupled lipid transmembrane transporter activity / ABC-type transporter activity / ATP hydrolysis activity / ATP binding / membrane Similarity search - Function | ||||||||||||
Biological species | ![]() ![]() ![]() ![]() | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.2 Å | ||||||||||||
![]() | Januliene D / Hofmann S / Medhdipour AR / Thomas C / Hummer G / Tampe R / Moeller A | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Conformation space of a heterodimeric ABC exporter under turnover conditions. Authors: Susanne Hofmann / Dovile Januliene / Ahmad R Mehdipour / Christoph Thomas / Erich Stefan / Stefan Brüchert / Benedikt T Kuhn / Eric R Geertsma / Gerhard Hummer / Robert Tampé / Arne Moeller / ![]() Abstract: Cryo-electron microscopy (cryo-EM) has the capacity to capture molecular machines in action. ATP-binding cassette (ABC) exporters are highly dynamic membrane proteins that extrude a wide range of ...Cryo-electron microscopy (cryo-EM) has the capacity to capture molecular machines in action. ATP-binding cassette (ABC) exporters are highly dynamic membrane proteins that extrude a wide range of substances from the cytosol and thereby contribute to essential cellular processes, adaptive immunity and multidrug resistance. Despite their importance, the coupling of nucleotide binding, hydrolysis and release to the conformational dynamics of these proteins remains poorly resolved, especially for heterodimeric and/or asymmetric ABC exporters that are abundant in humans. Here we present eight high-resolution cryo-EM structures that delineate the full functional cycle of an asymmetric ABC exporter in a lipid environment. Cryo-EM analysis under active turnover conditions reveals distinct inward-facing (IF) conformations-one of them with a bound peptide substrate-and previously undescribed asymmetric post-hydrolysis states with dimerized nucleotide-binding domains and a closed extracellular gate. By decreasing the rate of ATP hydrolysis, we could capture an outward-facing (OF) open conformation-an otherwise transient state vulnerable to substrate re-entry. The ATP-bound pre-hydrolysis and vanadate-trapped states are conformationally equivalent; both comprise co-existing OF conformations with open and closed extracellular gates. By contrast, the post-hydrolysis states from the turnover experiment exhibit asymmetric ATP and ADP occlusion after phosphate release from the canonical site and display a progressive separation of the nucleotide-binding domains and unlocking of the intracellular gate. Our findings reveal that phosphate release, not ATP hydrolysis, triggers the return of the exporter to the IF conformation. By mapping the conformational landscape during active turnover, aided by mutational and chemical modulation of kinetic rates to trap the key intermediates, we resolved fundamental steps of the substrate translocation cycle of asymmetric ABC transporters. | ||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 4 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 22 KB 22 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 9.2 KB | Display | ![]() |
Images | ![]() | 42.8 KB | ||
Masks | ![]() | 64 MB | ![]() | |
Others | ![]() ![]() | 59.5 MB 59.5 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6ragMC ![]() 4773C ![]() 4775C ![]() 4776C ![]() 4777C ![]() 4778C ![]() 4779C ![]() 4780C ![]() 4781C ![]() 6rafC ![]() 6rahC ![]() 6raiC ![]() 6rajC ![]() 6rakC ![]() 6ralC ![]() 6ramC ![]() 6ranC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | TmrAB in IF wide conformation under turnover conditions | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.077 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Mask #1
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Density Histograms |
-Half map: None
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Annotation | None | ||||||||||||
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Density Histograms |
-Half map: None
File | emd_4774_half_map_2.map | ||||||||||||
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Annotation | None | ||||||||||||
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Density Histograms |
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Sample components
-Entire : TmrAB in inward-facing wide conformation
Entire | Name: TmrAB in inward-facing wide conformation |
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Components |
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-Supramolecule #1: TmrAB in inward-facing wide conformation
Supramolecule | Name: TmrAB in inward-facing wide conformation / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Molecular weight | Theoretical: 150 KDa |
-Supramolecule #2: TmrAB
Supramolecule | Name: TmrAB / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#2 |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Supramolecule #3: nanobody
Supramolecule | Name: nanobody / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #3 |
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Source (natural) | Organism: ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Macromolecule #1: Multidrug resistance ABC transporter ATP-binding and permease protein
Macromolecule | Name: Multidrug resistance ABC transporter ATP-binding and permease protein type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 70.664797 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MTEDTYSKAF DRALFARILR YVWPYRLQVV LALLFLLVVT LAAAATPLFF KWAIDLALVP TEPRPLAERF HLLLWISLGF LAVRAVHFA ATYGETYLIQ WVGQRVLFDL RSDLFAKLMR LHPGFYDRNP VGRLMTRVTS DVDAINQFIT GGLVGVIADL F TLVGLLGF ...String: MTEDTYSKAF DRALFARILR YVWPYRLQVV LALLFLLVVT LAAAATPLFF KWAIDLALVP TEPRPLAERF HLLLWISLGF LAVRAVHFA ATYGETYLIQ WVGQRVLFDL RSDLFAKLMR LHPGFYDRNP VGRLMTRVTS DVDAINQFIT GGLVGVIADL F TLVGLLGF MLFLSPKLTL VVLLVAPVLL AVTTWVRLGM RSAYREMRLR LARVNAALQE NLSGVETIQL FVKEREREEK FD RLNRDLF RAWVEIIRWF ALFFPVVGFL GDFAVASLVY YGGGEVVRGA VSLGLLVAFV DYTRQLFQPL QDLSDKFNLF QGA MASAER IFGVLDTEEE LKDPEDPTPI RGFRGEVEFR DVWLAYTPKG VEPTEKDWVL KGVSFRVRPG EKVALVGATG AGKT SVVSL IARFYDPQRG CVFLDGVDVR RYRQEELRRH VGIVLQEPFL FSGTVLDNLR LFDPSVPPER VEEVARFLGA HEFIL RLPK GYQTVLGERG AGLSTGEKQL LALVRALLAS PDILLILDEA TASVDSETEK RLQEALYKAM EGRTSLIIAH RLSTIR HVD RILVFRKGRL VEEGSHEELL AKGGYYAALY RLQFQEAKLG GGGENLYFQG HHHHHHHHHH |
-Macromolecule #2: Multidrug resistance ABC transporter ATP-binding and permease protein
Macromolecule | Name: Multidrug resistance ABC transporter ATP-binding and permease protein type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 64.634457 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MTGRSAAPLL RRLWPYVGRY RWRYLWAVLA GLVSIFFFVL TPYFLRLAVD AVQAGRGFGV YALAIVASAA LSGLLSYAMR RLAVVASRQ VEYDLRRDLL HHLLTLDRDF YHKHRVGDLM NRLNTDLSAV REMVGPGILM GSRLSFLVLL AFLSMYAVNA R LAFYLTLI ...String: MTGRSAAPLL RRLWPYVGRY RWRYLWAVLA GLVSIFFFVL TPYFLRLAVD AVQAGRGFGV YALAIVASAA LSGLLSYAMR RLAVVASRQ VEYDLRRDLL HHLLTLDRDF YHKHRVGDLM NRLNTDLSAV REMVGPGILM GSRLSFLVLL AFLSMYAVNA R LAFYLTLI LPGIFLAMRF LLRLIDRRYR EAQEVFDRIS TLAQEAFSGI RVVKGYALER RMVAWFQDLN RLYVEKSLAL AR VEGPLHA LLGFLMGFAF LTVLWAGGAM VVRGELSVGE LVQFNAYLAQ LTWPILGLGW VMALYQRGLT SLRRLFELLD EKP AIRDED PLPLALEDLS GEVRFEGVGL KRDGRWLLRG LTLTIPEGMT LGITGRTGSG KSLLAALVPR LLDPSEGRVY VGGH EARRI PLAVLRKAVG VAPQEPFLFS ETILENIAFG LDEVDRERVE WAARLAGIHE EILAFPKGYE TVLGERGITL SGGQR QRVA LARALAKRPK ILILDDALSA VDAETEARIL QGLKTVLGKQ TTLLISHRTA ALRHADWIIV LDGGRIVEEG THESLL QAG GLYAEMDRLQ KEVEA |
-Macromolecule #3: Nanobody Nb9F10
Macromolecule | Name: Nanobody Nb9F10 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 14.594405 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MAQLQLVESG GGLVQPGDSL RLSCAVSGSA LDYNAIGWFR QAPGKEREGV ACISKITGNT AYADSVKGRF TISRDNAKNT VHLQMNSLK PEDTAVYYCA TVTAVLLPGR CVPGKYWGQG TPVTVSSHHH HHHEPEA |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #5: ADENOSINE-5'-DIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 5 / Number of copies: 1 / Formula: ADP |
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Molecular weight | Theoretical: 427.201 Da |
Chemical component information | ![]() ChemComp-ADP: |
-Macromolecule #6: ADENOSINE-5'-TRIPHOSPHATE
Macromolecule | Name: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 6 / Number of copies: 1 / Formula: ATP |
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Molecular weight | Theoretical: 507.181 Da |
Chemical component information | ![]() ChemComp-ATP: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 1 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: Quantifoil, UltrAuFoil / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Average exposure time: 8.0 sec. / Average electron dose: 62.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |