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- PDB-8the: Cryo-EM structure of Pseudomonas aeruginosa TonB-dependent transp... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8the | ||||||
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Title | Cryo-EM structure of Pseudomonas aeruginosa TonB-dependent transporter PhuR in complex with synthetic antibody and heme | ||||||
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Function / homology | ![]() heme transport / siderophore transmembrane transport / heme transmembrane transporter activity / siderophore uptake transmembrane transporter activity / outer membrane / cell outer membrane / cellular response to iron ion Similarity search - Function | ||||||
Biological species | ![]() ![]() ![]() ![]() ![]() | ||||||
Method | ![]() ![]() ![]() | ||||||
![]() | Knejski, P. / Erramilli, S.K. / Kossiakoff, A.A. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Chaperone-assisted cryo-EM structure of P. aeruginosa PhuR reveals molecular basis for heme binding. Authors: Paweł P Knejski / Satchal K Erramilli / Anthony A Kossiakoff / ![]() ![]() Abstract: Pathogenic bacteria, such as Pseudomonas aeruginosa, depend on scavenging heme for the acquisition of iron, an essential nutrient. The TonB-dependent transporter (TBDT) PhuR is the major heme uptake ...Pathogenic bacteria, such as Pseudomonas aeruginosa, depend on scavenging heme for the acquisition of iron, an essential nutrient. The TonB-dependent transporter (TBDT) PhuR is the major heme uptake protein in P. aeruginosa clinical isolates. However, a comprehensive understanding of heme recognition and TBDT transport mechanisms, especially PhuR, remains limited. In this study, we employed single-particle cryogenic electron microscopy (cryo-EM) and a phage display-generated synthetic antibody (sAB) as a fiducial marker to enable the determination of a high-resolution (2.5 Å) structure of PhuR with a bound heme. Notably, the structure reveals iron coordination by Y529 on a conserved extracellular loop, shedding light on the role of tyrosine in heme binding. Biochemical assays and negative-stain EM demonstrated that the sAB specifically targets the heme-bound state of PhuR. These findings provide insights into PhuR's heme binding and offer a template for developing conformation-specific sABs against outer membrane proteins (OMPs) for structure-function investigations. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 212.1 KB | Display | ![]() |
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PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 41255MC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
Experimental dataset #1 | Data reference: ![]() |
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Links
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Assembly
Deposited unit | ![]()
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1 |
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Components
-Protein , 1 types, 1 molecules A
#1: Protein | Mass: 86370.320 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() ![]() ![]() |
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-Antibody , 2 types, 2 molecules HL
#2: Antibody | ![]() Mass: 25525.445 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() ![]() |
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#3: Antibody | ![]() Mass: 23212.715 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() ![]() ![]() |
-Non-polymers , 3 types, 83 molecules ![](data/chem/img/HEM.gif)
![](data/chem/img/CTQ.gif)
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#4: Chemical | ChemComp-HEM / ![]() |
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#5: Chemical | ChemComp-CTQ / ( |
#6: Water | ChemComp-HOH / ![]() |
-Details
Has ligand of interest | Y |
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-Experimental details
-Experiment
Experiment | Method: ![]() |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: ![]() |
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Sample preparation
Component | Name: Heme bound P. aeruginosa PhuR with bound synthetic antibody Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: ![]() ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() ![]() |
Buffer solution | pH: 7.5 |
Specimen | Conc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied![]() ![]() |
Specimen support | 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 / Humidity: 100 % / Chamber temperature: 281 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source![]() ![]() |
Electron lens | Mode: BRIGHT FIELD![]() |
Image recording | Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of real images: 5031 |
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Processing
EM software |
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CTF correction![]() | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry![]() | ||||||||||||||||||||||||||||||||
3D reconstruction![]() | Resolution: 2.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 292512 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building | Source name: AlphaFold / Type: in silico model |