|Entry||Database: EMDB / ID: EMD-9832|
|Title||Tetrameric PepTSo2 incorporated in salipro nano particle|
|Sample||Tetrameric PepTSo2 with saposin A|
|Function / homology|
Function and homology information
tripeptide transmembrane transporter activity / peptide:proton symporter activity / oligopeptide transmembrane transporter activity / dipeptide transmembrane transporter activity / peptide transmembrane transporter activity / integral component of plasma membrane / integral component of membrane / identical protein binding / plasma membrane
Dipeptide/tripeptide permease / Proton-dependent oligopeptide transporter family / Major facilitator superfamily domain / MFS transporter superfamily / POT family
Proton:oligopeptide symporter POT family
|Source||Shewanella oneidensis (bacteria) / Shewanella oneidensis MR-1 (bacteria)|
|Method||single particle reconstruction / cryo EM / Resolution: 4.1 Å|
|Authors||Kawamoto A / Matoba K / Takagi J|
|Citation||Journal: Acta Crystallogr F Struct Biol Commun / Year: 2019|
Title: Structural basis for oligomerization of the prokaryotic peptide transporter PepT.
Authors: Reina Nagamura / Masahiro Fukuda / Akihiro Kawamoto / Kyoko Matoba / Naoshi Dohmae / Ryuichiro Ishitani / Junichi Takagi / Osamu Nureki /
Abstract: Proton-dependent oligopeptide transporters (POTs) belong to the major facilitator superfamily (MFS) and transport dipeptides and tripeptides from the extracellular environment into the target cell. ...Proton-dependent oligopeptide transporters (POTs) belong to the major facilitator superfamily (MFS) and transport dipeptides and tripeptides from the extracellular environment into the target cell. The human POTs PepT1 and PepT2 are also involved in the absorption of various orally ingested drugs. Previously reported structures revealed that the bacterial POTs possess 14 helices, of which H1-H6 and H7-H12 constitute the typical MFS fold and the residual two helices are involved in the cytoplasmic linker. PepT from Shewanella oneidensis is a unique POT which reportedly assembles as a 200 kDa tetramer. Although the previously reported structures suggested the importance of H12 for tetramer formation, the structural basis for the PepT-specific oligomerization remains unclear owing to the lack of a high-resolution tetrameric structure. In this study, the expression and purification conditions for tetrameric PepT were optimized. A single-particle cryo-EM analysis revealed the tetrameric structure of PepT incorporated into Salipro nanoparticles at 4.1 Å resolution. Furthermore, a combination of lipidic cubic phase (LCP) crystallization and an automated data-processing system for multiple microcrystals enabled crystal structures of PepT to be determined at resolutions of 3.5 and 3.9 Å. The present structures in a lipid bilayer revealed the detailed mechanism for the tetrameric assembly of PepT, in which a characteristic extracellular loop (ECL) interacts with two asparagine residues on H12 which were reported to be important for tetramerization and plays an essential role in oligomeric assembly. This study provides valuable insights into the oligomerization mechanism of this MFS-type transporter, which will further pave the way for understanding other oligomeric membrane proteins.
|Validation Report||PDB-ID: 6ji1|
SummaryFull reportAbout validation report
|Date||Deposition: Feb 19, 2019 / Header (metadata) release: May 15, 2019 / Map release: May 15, 2019 / Update: May 15, 2019|
|Structure viewer||EM map: |
Downloads & links
|File||Download / File: emd_9832.map.gz / Format: CCP4 / Size: 107.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 0.87 Å|
|Symmetry||Space group: 1|
CCP4 map header:
-Entire Tetrameric PepTSo2 with saposin A
|Entire||Name: Tetrameric PepTSo2 with saposin A / Number of components: 2|
-Component #1: protein, Tetrameric PepTSo2 with saposin A
|Protein||Name: Tetrameric PepTSo2 with saposin A / Recombinant expression: No|
|Source||Species: Shewanella oneidensis (bacteria)|
|Source (engineered)||Expression System: Escherichia coli K-12 (bacteria) / Strain: K-12|
-Component #2: protein, Proton:oligopeptide symporter POT family
|Protein||Name: Proton:oligopeptide symporter POT family / Number of Copies: 4 / Recombinant expression: No|
|Mass||Theoretical: 57.626559 kDa|
|Source||Species: Shewanella oneidensis MR-1 (bacteria) / Strain: MR-1|
|Source (engineered)||Expression System: Escherichia coli K-12 (bacteria)|
|Specimen||Specimen state: Particle / Method: cryo EM|
|Sample solution||Specimen conc.: 0.5 mg/mL / pH: 7.4|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 100 % / Details: blotted for 4.5 seconds before plunging.|
-Electron microscopy imaging
Model: Talos Arctica / Image courtesy: FEI Company
|Imaging||Microscope: FEI TALOS ARCTICA|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 82 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 96000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1000.0 - 3000.0 nm|
|Specimen Holder||Model: OTHER / Temperature: (79.55 - 79.55 K)|
|Camera||Detector: FEI FALCON III (4k x 4k)|
|Image acquisition||Number of digital images: 1609|
-Atomic model buiding
|Modeling #1||Refinement protocol: flexible / Refinement space: REAL|
-Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)
EMDB accession codes are about to change! (news from PDBe EMDB page)
- The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force. (see PDBe EMDB page)
- The EM Navigator/Yorodumi systems omit the EMD- prefix.
Related info.: Q: What is "EMD"? / ID/Accession-code notation in Yorodumi/EM Navigator
-Jul 12, 2017. Major update of PDB
Major update of PDB
- wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary. This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated. See below links for details.
- In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software). Now, EM Navigator and Yorodumi are based on the updated data.
+Jun 16, 2017. Omokage search with filter
Omokage search with filter
- Result of Omokage search can be filtered by keywords and the database types
Related info.: Omokage search
+Sep 15, 2016. EM Navigator & Yorodumi renewed
EM Navigator & Yorodumi renewed
- New versions of EM Navigator and Yorodumi started
Related info.: Changes in new EM Navigator and Yorodumi
+Aug 31, 2016. New EM Navigator & Yorodumi
New EM Navigator & Yorodumi
- In 15th Sep 2016, the development versions of EM Navigator and Yorodumi will replace the official versions.
- Current version will continue as 'legacy version' for some time.
Related info.: Changes in new EM Navigator and Yorodumi / EM Navigator / Yorodumi
Thousand views of thousand structures
- Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
- This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
Related info.: EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi