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- PDB-8y5f: Cryo-EM structure of E.coli spermidine transporter PotABC -

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Basic information

Entry
Database: PDB / ID: 8y5f
TitleCryo-EM structure of E.coli spermidine transporter PotABC
Components
  • Spermidine/putrescine import ATP-binding protein PotA
  • Spermidine/putrescine transport system permease protein PotB
  • Spermidine/putrescine transport system permease protein PotC
KeywordsTRANSPORT PROTEIN / ABC transporter
Function / homology
Function and homology information


ABC-type polyamine transporter / ABC-type putrescine transporter activity / spermidine transmembrane transport / putrescine transport / ABC-type polyamine transporter activity / ATP-binding cassette (ABC) transporter complex / nucleotide binding / ATP hydrolysis activity / ATP binding / membrane / plasma membrane
Similarity search - Function
Spermidine/putrescine import ATP-binding protein potA family profile. / Spermidine/putrescine ABC transporter, ATP-binding subunit / PotA, ATP-binding domain / : / : / Transport-associated OB, type 2 / TOBE domain / Molybdate/tungstate binding, C-terminal / ABC transporter type 1, transmembrane domain MetI-like / MetI-like superfamily ...Spermidine/putrescine import ATP-binding protein potA family profile. / Spermidine/putrescine ABC transporter, ATP-binding subunit / PotA, ATP-binding domain / : / : / Transport-associated OB, type 2 / TOBE domain / Molybdate/tungstate binding, C-terminal / ABC transporter type 1, transmembrane domain MetI-like / MetI-like superfamily / Binding-protein-dependent transport system inner membrane component / ABC transporter integral membrane type-1 domain profile. / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Spermidine/putrescine transport system permease protein PotC / Spermidine/putrescine import ATP-binding protein PotA
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.13 Å
AuthorsQiao, Z. / Gao, Y.G.
Funding support Singapore, 1items
OrganizationGrant numberCountry
Ministry of Education (MoE, Singapore) Singapore
CitationJournal: Sci Adv / Year: 2024
Title: Structural insights into polyamine spermidine uptake by the ABC transporter PotD-PotABC.
Authors: Zhu Qiao / Phong Hoa Do / Joshua Yi Yeo / Rya Ero / Zhuowen Li / Liying Zhan / Sandip Basak / Yong-Gui Gao /
Abstract: Polyamines, characterized by their polycationic nature, are ubiquitously present in all organisms and play numerous cellular functions. Among polyamines, spermidine stands out as the predominant type ...Polyamines, characterized by their polycationic nature, are ubiquitously present in all organisms and play numerous cellular functions. Among polyamines, spermidine stands out as the predominant type in both prokaryotic and eukaryotic cells. The PotD-PotABC protein complex in , belonging to the adenosine triphosphate-binding cassette transporter family, is a spermidine-preferential uptake system. Here, we report structural details of the polyamine uptake system PotD-PotABC in various states. Our analyses reveal distinct "inward-facing" and "outward-facing" conformations of the PotD-PotABC transporter, as well as conformational changes in the "gating" residues (F222, Y223, D226, and K241 in PotB; Y219 and K223 in PotC) controlling spermidine uptake. Therefore, our structural analysis provides insights into how the PotD-PotABC importer recognizes the substrate-binding protein PotD and elucidates molecular insights into the spermidine uptake mechanism of bacteria.
History
DepositionJan 31, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Oct 9, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Spermidine/putrescine import ATP-binding protein PotA
B: Spermidine/putrescine transport system permease protein PotB
C: Spermidine/putrescine transport system permease protein PotC
D: Spermidine/putrescine import ATP-binding protein PotA


Theoretical massNumber of molelcules
Total (without water)147,5254
Polymers147,5254
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Spermidine/putrescine import ATP-binding protein PotA


Mass: 43079.137 Da / Num. of mol.: 2 / Mutation: E173Q
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: potA, b1126, JW1112 / Production host: Escherichia coli (E. coli) / References: UniProt: P69874, ABC-type polyamine transporter
#2: Protein Spermidine/putrescine transport system permease protein PotB


Mass: 32234.164 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: potB, b1125, JW1111 / Production host: Escherichia coli (E. coli)
#3: Protein Spermidine/putrescine transport system permease protein PotC


Mass: 29132.850 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: potC, b1124, JW1110 / Production host: Escherichia coli (E. coli) / References: UniProt: P0AFK6
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: ABC transporter / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 147.3 kDa/nm / Experimental value: YES
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenConc.: 10 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1000 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.18.2_3874: / Category: model refinement
CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 3.13 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 90050 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00610037
ELECTRON MICROSCOPYf_angle_d0.68513609
ELECTRON MICROSCOPYf_dihedral_angle_d8.5911336
ELECTRON MICROSCOPYf_chiral_restr0.0461597
ELECTRON MICROSCOPYf_plane_restr0.0061721

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