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Yorodumi- PDB-7nnl: Cryo-EM structure of the KdpFABC complex in an E1-ATP conformatio... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7nnl | ||||||||||||||||||
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Title | Cryo-EM structure of the KdpFABC complex in an E1-ATP conformation loaded with K+ | ||||||||||||||||||
Components | (Potassium-transporting ATPase ...) x 4 | ||||||||||||||||||
Keywords | MEMBRANE PROTEIN / P-type ATPase / superfamily of K+ transporters (SKT) / potassium uptake system / ATP analogue | ||||||||||||||||||
Function / homology | Function and homology information P-type K+ transporter / P-type potassium transmembrane transporter activity / potassium:proton antiporter complex / potassium ion-transporting ATPase complex / plasma membrane => GO:0005886 / monoatomic cation transmembrane transport / potassium ion binding / potassium ion transmembrane transport / potassium ion transport / hydrolase activity ...P-type K+ transporter / P-type potassium transmembrane transporter activity / potassium:proton antiporter complex / potassium ion-transporting ATPase complex / plasma membrane => GO:0005886 / monoatomic cation transmembrane transport / potassium ion binding / potassium ion transmembrane transport / potassium ion transport / hydrolase activity / magnesium ion binding / ATP hydrolysis activity / ATP binding / plasma membrane Similarity search - Function | ||||||||||||||||||
Biological species | Escherichia coli (E. coli) | ||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å | ||||||||||||||||||
Authors | Silberberg, J.M. / Corey, R.A. / Hielkema, L. / Stock, C. / Stansfeld, P.J. / Paulino, C. / Haenelt, I. | ||||||||||||||||||
Funding support | Netherlands, Germany, United Kingdom, 5items
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Citation | Journal: Nat Commun / Year: 2021 Title: Deciphering ion transport and ATPase coupling in the intersubunit tunnel of KdpFABC. Authors: Jakob M Silberberg / Robin A Corey / Lisa Hielkema / Charlott Stock / Phillip J Stansfeld / Cristina Paulino / Inga Hänelt / Abstract: KdpFABC, a high-affinity K pump, combines the ion channel KdpA and the P-type ATPase KdpB to secure survival at K limitation. Here, we apply a combination of cryo-EM, biochemical assays, and MD ...KdpFABC, a high-affinity K pump, combines the ion channel KdpA and the P-type ATPase KdpB to secure survival at K limitation. Here, we apply a combination of cryo-EM, biochemical assays, and MD simulations to illuminate the mechanisms underlying transport and the coupling to ATP hydrolysis. We show that ions are transported via an intersubunit tunnel through KdpA and KdpB. At the subunit interface, the tunnel is constricted by a phenylalanine, which, by polarized cation-π stacking, controls K entry into the canonical substrate binding site (CBS) of KdpB. Within the CBS, ATPase coupling is mediated by the charge distribution between an aspartate and a lysine. Interestingly, individual elements of the ion translocation mechanism of KdpFABC identified here are conserved among a wide variety of P-type ATPases from different families. This leads us to the hypothesis that KdpB might represent an early descendant of a common ancestor of cation pumps. #1: Journal: Biorxiv / Year: 2021 Title: Deciphering ion transport and ATPase coupling in the intersubunit tunnel of KdpFABC Authors: Silberberg, J.M. / Corey, R.A. / Hielkema, L. / Stock, C. / Stansfeld, P.J. / Paulino, C. / Hanelt, I. | ||||||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7nnl.cif.gz | 251.8 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7nnl.ent.gz | 207.6 KB | Display | PDB format |
PDBx/mmJSON format | 7nnl.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7nnl_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
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Full document | 7nnl_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 7nnl_validation.xml.gz | 53 KB | Display | |
Data in CIF | 7nnl_validation.cif.gz | 79.7 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/nn/7nnl ftp://data.pdbj.org/pub/pdb/validation_reports/nn/7nnl | HTTPS FTP |
-Related structure data
Related structure data | 12478MC 7nnpC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Potassium-transporting ATPase ... , 4 types, 4 molecules ACDB
#1: Protein | Mass: 59218.613 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: kdpA, D9J52_12825, SAMEA3752559_01047 / Production host: Escherichia coli (E. coli) / Strain (production host): LB2003 / References: UniProt: A0A2S5ZPF1 |
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#2: Protein | Mass: 20281.035 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) Gene: kdpC, A9R57_11470, ACU57_12815, AM464_20755, AUQ13_21370, BANRA_01742, BANRA_03870, BHS87_03640, BJJ90_18865, BMA87_25495, BON98_02815, BUE81_20120, BvCms12BK_01599, BvCms2454_04067, ...Gene: kdpC, A9R57_11470, ACU57_12815, AM464_20755, AUQ13_21370, BANRA_01742, BANRA_03870, BHS87_03640, BJJ90_18865, BMA87_25495, BON98_02815, BUE81_20120, BvCms12BK_01599, BvCms2454_04067, BvCmsHHP001_00774, BvCmsHHP056_03395, BvCmsKSP026_01797, BW690_07920, C5F72_18220, C5F73_11285, C5N07_14650, C9E25_26865, CA593_25675, CG692_12945, CI694_20645, D0X26_17510, D2185_15885, D3821_21865, D3Y67_05220, D4718_15195, D9C99_19315, D9D20_12860, D9D44_15890, D9G69_25005, D9J52_12835, D9Z28_23025, DAH34_18920, DBQ99_18020, DJ503_06890, DL326_22940, DNC98_20170, DT034_24065, DTL43_19970, DXT73_03320, E2119_26465, E2134_18585, E2135_05100, E4K51_13735, E4K53_14000, E4K55_14005, E4K61_12665, E5P28_21685, EA213_10645, EAI52_22450, EC3234A_8c00230, ECTO6_03352, ED307_21640, EEP23_23705, EG808_19380, EI021_12890, EI028_12880, EI032_09320, EI041_12810, EIZ93_03125, EL75_3091, EL79_3184, EL80_5457, ERS085365_04786, ERS085416_02369, ERS139211_04506, ERS150873_01994, EYD11_15990, EYV18_15885, F1E19_24880, F9040_19740, FNW97_03505, FRV13_05295, FV293_12595, G5688_17415, GII66_07600, GKF39_08170, GKF74_06650, GKF86_08095, GKF89_05805, GP689_15935, GP954_16115, GP979_23630, GQE33_23970, GQE34_25250, GQE64_10530, GQM06_27255, GQM17_23755, GRW05_21650, GRW42_11025, GRW57_15570, GRW80_11135, GRW81_16520, HVY93_16490, HX136_18175, NCTC10963_03490, NCTC8500_03902, NCTC9045_04050, NCTC9058_03392, NCTC9062_04759, NCTC9073_02588, NCTC9706_00826, PGD_02635, RK56_025175, SAMEA3472044_02479, SAMEA3472047_02083, SAMEA3472080_00272, SAMEA3484427_03080, SAMEA3484429_03302, SAMEA3753097_03043, SAMEA3753300_04144, WP2S18E08_32510, WP7S17E04_30600 Production host: Escherichia coli (E. coli) / Strain (production host): LB2003 / References: UniProt: A0A037YI39 |
#3: Protein/peptide | Mass: 2853.463 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli (E. coli) / Gene: kdpF, b4513, JW0687 / Production host: Escherichia coli (E. coli) / Strain (production host): LB2003 / References: UniProt: P36937 |
#4: Protein | Mass: 72346.859 Da / Num. of mol.: 1 / Mutation: D307N Source method: isolated from a genetically manipulated source Details: S162-P / Source: (gene. exp.) Escherichia coli (E. coli) Gene: kdpB, ACU57_12810, AUQ13_21365, BANRA_01741, BANRA_03871, BJJ90_18860, BMA87_25490, BUE81_20115, BvCms12BK_01600, BvCms2454_04068, BvCmsHHP001_00773, BvCmsKSP026_01798, BW690_07915, C5F73_ ...Gene: kdpB, ACU57_12810, AUQ13_21365, BANRA_01741, BANRA_03871, BJJ90_18860, BMA87_25490, BUE81_20115, BvCms12BK_01600, BvCms2454_04068, BvCmsHHP001_00773, BvCmsKSP026_01798, BW690_07915, C5F73_11290, C5N07_14655, C9E25_26860, CA593_25670, CG692_12950, D0X26_17505, D2185_15880, D3821_21860, D3Y67_05225, D9C99_19310, D9D44_15885, D9G69_25000, D9J52_12830, D9Z28_23020, DBQ99_18015, DJ503_06885, DL326_22935, DNC98_20165, DT034_24060, DTL43_19975, DXT73_03325, E2119_26470, E2134_18590, E2135_05105, E4K51_13740, E4K53_14005, E4K55_14010, E4K61_12660, EA213_10640, EAI52_22455, EC3234A_8c00240, ECTO6_03351, ED307_21635, EEP23_23700, EG808_19375, EI021_12885, EI028_12885, EI041_12805, EPT01_08650, ERS085365_04785, ERS085416_02368, ERS139211_04507, ERS150873_01995, EXX71_13725, EYD11_15985, EYV18_15890, F1E19_24885, FNW97_03510, FV293_12590, FWK02_15235, GKF39_08175, GKF74_06655, GKF86_08100, GKF89_05800, GP689_15940, GQE33_23975, GQE34_25255, GQE64_10535, GQM17_23760, GRW05_21645, GRW42_11030, GRW80_11130, GRW81_16515, HVY93_16485, HX136_18170, NCTC8500_03901, NCTC9045_04049, NCTC9062_04760, PGD_02634, RK56_025170, SAMEA3472047_02084, SAMEA3484427_03079, SAMEA3484429_03303, SAMEA3753300_04145, WP2S18E08_32500, WP7S17E04_30590 Production host: Escherichia coli (E. coli) / Strain (production host): LB2003 / References: UniProt: A0A024L5I2, P-type K+ transporter |
-Non-polymers , 3 types, 13 molecules
#5: Chemical | ChemComp-K / #6: Chemical | #7: Chemical | ChemComp-ACP / | |
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-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: KdpFABC / Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT |
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Molecular weight | Value: 0.157 MDa / Experimental value: NO |
Source (natural) | Organism: Escherichia coli (E. coli) |
Source (recombinant) | Organism: Escherichia coli (E. coli) / Strain: LB2003 |
Buffer solution | pH: 8 Details: 10 mM Tris-HCl pH 8, 10 mM MgCl2, 10 mM NaCl and 0.0125% DDM |
Specimen | Conc.: 3.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: at 5 mA / Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 293 K |
-Electron microscopy imaging
Experimental equipment | Model: Talos Arctica / Image courtesy: FEI Company |
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Microscopy | Model: FEI TALOS ARCTICA |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 49407 X / Calibrated magnification: 49407 X / Nominal defocus max: 2000 nm / Nominal defocus min: 500 nm / Calibrated defocus min: 500 nm / Calibrated defocus max: 2000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 105 K / Temperature (min): 90 K |
Image recording | Average exposure time: 9 sec. / Electron dose: 52 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 6 / Num. of real images: 5831 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Width: 3838 / Height: 3710 / Movie frames/image: 60 / Used frames/image: 1-60 |
-Processing
Software | Name: PHENIX / Version: 1.19.1_4122: / Classification: refinement | ||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 331673 | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 160776 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building | Space: REAL | ||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 6HRA | ||||||||||||||||||||||||||||||||
Refine LS restraints |
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