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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-23269 | ||||||||||||
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Title | Cryo-EM Structure of KdpFABC in E2-P state with BeF3 | ||||||||||||
![]() | non-linear refinement with cryoSPARC | ||||||||||||
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Function / homology | ![]() P-type K+ transporter / potassium:proton antiporter complex / potassium ion-transporting ATPase complex / P-type potassium transmembrane transporter activity / monoatomic cation transmembrane transport / ![]() ![]() ![]() ![]() ![]() ![]() Similarity search - Function | ||||||||||||
Biological species | ![]() ![]() ![]() | ||||||||||||
Method | ![]() ![]() | ||||||||||||
![]() | Sweet ME / Larsen C | ||||||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: Structural basis for potassium transport in prokaryotes by KdpFABC. Authors: Marie E Sweet / Casper Larsen / Xihui Zhang / Michael Schlame / Bjørn P Pedersen / David L Stokes / ![]() ![]() Abstract: KdpFABC is an oligomeric K transport complex in prokaryotes that maintains ionic homeostasis under stress conditions. The complex comprises a channel-like subunit (KdpA) from the superfamily of K ...KdpFABC is an oligomeric K transport complex in prokaryotes that maintains ionic homeostasis under stress conditions. The complex comprises a channel-like subunit (KdpA) from the superfamily of K transporters and a pump-like subunit (KdpB) from the superfamily of P-type ATPases. Recent structural work has defined the architecture and generated contradictory hypotheses for the transport mechanism. Here, we use substrate analogs to stabilize four key intermediates in the reaction cycle and determine the corresponding structures by cryogenic electron microscopy. We find that KdpB undergoes conformational changes consistent with other representatives from the P-type superfamily, whereas KdpA, KdpC, and KdpF remain static. We observe a series of spherical densities that we assign as K or water and which define a pathway for K transport. This pathway runs through an intramembrane tunnel in KdpA and delivers ions to sites in the membrane domain of KdpB. Our structures suggest a mechanism where ATP hydrolysis is coupled to K transfer between alternative sites in KdpB, ultimately reaching a low-affinity site where a water-filled pathway allows release of K to the cytoplasm. | ||||||||||||
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 | ![]() | 118.2 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 27.8 KB 27.8 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 11.2 KB | Display | ![]() |
Images | ![]() | 87.4 KB | ||
Masks | ![]() | 125 MB | ![]() | |
Filedesc metadata | ![]() | 8.1 KB | ||
Others | ![]() ![]() | 115.9 MB 115.9 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7lc6MC ![]() 7bgyC ![]() 7bh1C ![]() 7bh2C ![]() 7lc3C 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 | non-linear refinement with cryoSPARC | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.079 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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Projections & Slices |
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Density Histograms |
-Half map: Half mask A from cryoSPARC
File | emd_23269_half_map_1.map | ||||||||||||
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Annotation | Half mask A from cryoSPARC | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half mask B from cryoSPARC
File | emd_23269_half_map_2.map | ||||||||||||
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Annotation | Half mask B from cryoSPARC | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : KdpFABC in E2-P intermediate state with BeF3
Entire | Name: KdpFABC in E2-P intermediate state with BeF3 |
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Components |
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-Supramolecule #1: KdpFABC in E2-P intermediate state with BeF3
Supramolecule | Name: KdpFABC in E2-P intermediate state with BeF3 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 Details: KdpFABC complex solublized in decyl maltopyranoside (DM) detergent. |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 160 KDa |
-Macromolecule #1: Potassium-transporting ATPase potassium-binding subunit
Macromolecule | Name: Potassium-transporting ATPase potassium-binding subunit type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 59.247684 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: MAAQGFLLIA TFLLVLMVLA RPLGSGLARL INDIPLPGTT GVERVLFRAL GVSDREMNWK QYLCAILGLN MLGLAVLFFM LLGQHYLPL NPQQLPGLSW DLALNTAVSF VTNTNWRSYS GETTLSYFSQ MAGLTVQNFL SAASGIAVIF ALIRAFTRQS M STLGNAWV ...String: MAAQGFLLIA TFLLVLMVLA RPLGSGLARL INDIPLPGTT GVERVLFRAL GVSDREMNWK QYLCAILGLN MLGLAVLFFM LLGQHYLPL NPQQLPGLSW DLALNTAVSF VTNTNWRSYS GETTLSYFSQ MAGLTVQNFL SAASGIAVIF ALIRAFTRQS M STLGNAWV DLLRITLWVL VPVALLIALF FIQQGALQNF LPYQAVNTVE GAQQLLPMGP VASQEAIKML GTNGGGFFNA NS SHPFENP TALTNFVQML AIFLIPTALC FAFGEVMGDR RQGRMLLWAM SVIFVICVGV VMWAEVQGNP HLLALGTDSS INM EGKESR FGVLVSSLFA VVTTAASCGA VIAMHDSFTA LGGMVPMWLM QIGEVVFGGV GSGLYGMMLF VLLAVFIAGL MIGR TPEYL GKKIDVREMK LTALAILVTP TLVLMGAALA MMTDAGRSAM LNPGPHGFSE VLYAVSSAAN NNGSAFAGLS ANSPF WNCL LAFCMFVGRF GVIIPVMAIA GSLVSKKSQA ASSGTLPTHG PLFVGLLIGT VLLVGALTFI PALALGPVAE YLS UniProtKB: Potassium-transporting ATPase potassium-binding subunit |
-Macromolecule #2: Potassium-transporting ATPase ATP-binding subunit
Macromolecule | Name: Potassium-transporting ATPase ATP-binding subunit / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: P-type K+ transporter |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 72.251867 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: MSRKQLALFE PTLVVQALKE AVKKLNPQAQ WRNPVMFIVW IGSLLTTCIS IAMASGAMPG NALFSAAISG WLWITVLFAN FAEALAEGR SKAQANSLKG VKKTAFARKL REPKYGAAAD KVPADQLRKG DIVLVEAGDI IPCDGEVIEG GASVDESAIT G EAAPVIRE ...String: MSRKQLALFE PTLVVQALKE AVKKLNPQAQ WRNPVMFIVW IGSLLTTCIS IAMASGAMPG NALFSAAISG WLWITVLFAN FAEALAEGR SKAQANSLKG VKKTAFARKL REPKYGAAAD KVPADQLRKG DIVLVEAGDI IPCDGEVIEG GASVDESAIT G EAAPVIRE SGGDFASVTG GTRILSDWLV IECSVNPGET FLDRMIAMVE GAQRRKTPNE IALTILLIAL TIVFLLATAT LW PFSAWGG NAVSVTVLVA LLVCLIPTTI GGLLSAIGVA GMSRMLGANV IATSGRAVEA AGDVDVLLLD KTGTITLGNR QAS EFIPAQ GVDEKTLADA AQLASLADET PEGRSIVILA KQRFNLRERD VQSLHATFVP FTAQSRMSGI NIDNRMIRKG SVDA IRRHV EANGGHFPTD VDQKVDQVAR QGATPLVVVE GSRVLGVIAL KDIVKGGIKE RFAQLRKMGI KTVMITGDNR LTAAA IAAE AGVDDFLAEA TPEAKLALIR QYQAEGRLVA MTGDGTNDAP ALAQADVAVA MNSGTQAAKE AGNMVDLDSN PTKLIE VVH IGKQMLMTRG SLTTFSIAND VAKYFAIIPA AFAATYPQLN ALNIMCLHSP DSAILSAVIF NALIIVFLIP LALKGVS YK PLTASAMLRR NLWIYGLGGL LVPFIGIKVI DLLLTVCGLV UniProtKB: Potassium-transporting ATPase ATP-binding subunit |
-Macromolecule #3: Potassium-transporting ATPase KdpC subunit
Macromolecule | Name: Potassium-transporting ATPase KdpC subunit / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 22.299225 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: MSGLRPALST FIFLLLITGG VYPLLTTVLG QWWFPWQANG SLIREGDTVR GSALIGQNFT GNGYFHGRPS ATAEMPYNPQ ASGGSNLAV SNPELDKLIA ARVAALRAAN PDASASVPVE LVTASASGLD NNITPQAAAW QIPRVAKARN LSVEQLTQLI A KYSQQPLV ...String: MSGLRPALST FIFLLLITGG VYPLLTTVLG QWWFPWQANG SLIREGDTVR GSALIGQNFT GNGYFHGRPS ATAEMPYNPQ ASGGSNLAV SNPELDKLIA ARVAALRAAN PDASASVPVE LVTASASGLD NNITPQAAAW QIPRVAKARN LSVEQLTQLI A KYSQQPLV KYIGQPVVNI VELNLALDKL DEGTGLVPRG SSHHHHHHHH UniProtKB: Potassium-transporting ATPase KdpC subunit |
-Macromolecule #4: Potassium-transporting ATPase KdpF subunit
Macromolecule | Name: Potassium-transporting ATPase KdpF subunit / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 3.071714 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: MSAGVITGVL LVFLLLGYLV YALINAEAF UniProtKB: Potassium-transporting ATPase KdpF subunit |
-Macromolecule #5: POTASSIUM ION
Macromolecule | Name: POTASSIUM ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: K |
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Molecular weight | Theoretical: 39.098 Da |
-Macromolecule #6: (2R)-3-(((2-aminoethoxy)(hydroxy)phosphoryl)oxy)-2-(palmitoyloxy)...
Macromolecule | Name: (2R)-3-(((2-aminoethoxy)(hydroxy)phosphoryl)oxy)-2-(palmitoyloxy)propyl (E)-octadec-9-enoate type: ligand / ID: 6 / Number of copies: 1 / Formula: 9Y0 |
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Molecular weight | Theoretical: 717.996 Da |
Chemical component information | ![]() ChemComp-9Y0: |
-Macromolecule #7: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 7 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #8: water
Macromolecule | Name: water / type: ligand / ID: 8 / Number of copies: 4 / Formula: HOH |
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Molecular weight | Theoretical: 18.015 Da |
Chemical component information | ![]() ChemComp-HOH: |
-Experimental details
-Structure determination
Method | ![]() |
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Aggregation state | particle |
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Sample preparation
Concentration | 4.5 mg/mL | ||||||||||||||||||||||||
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Buffer | pH: 7.5 Component:
Details: The beryllium fluoride in the buffer was added to the protein mixture containing all other buffer components in the form of a pre-incubated mixture with the final concentrations 2.5 mM BeSO4 and 10 mM NaF. | ||||||||||||||||||||||||
Grid | Model: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Support film - Film thickness: 50 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: AIR Details: PELCO easiGlow Glow Discharge Cleaning System used for glow discharge with default settings. | ||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV Details: 3 uL of protein mixture applied to grid. Blot time 4 seconds, blot force 0, no wait before plunging.. | ||||||||||||||||||||||||
Details | DM-solublized KdpFABC complex purified through SEC the same day as grid freezing. Peak fraction diluted to 4.5 mg/mL and complexed with beryllium fluoride solution for >30 min at room temperature prior to application to grid. |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD![]() |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Temperature | Min: 77.0 K / Max: 77.0 K |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 2567 / Average exposure time: 2.5 sec. / Average electron dose: 50.0 e/Å2 |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |