[English] 日本語
Yorodumi
- PDB-8pd0: cryo-EM structure of Doa10 in MSP1E3D1 -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 8pd0
Titlecryo-EM structure of Doa10 in MSP1E3D1
ComponentsERAD-associated E3 ubiquitin-protein ligase DOA10
KeywordsLIGASE / ERAD / Doa10 / March6 / TEB4 / retrotranslocation / ubiquitination / Ubc6 / sybody / SQLE / squalenemonooxygenase
Function / homology
Function and homology information


Doa10p ubiquitin ligase complex / nuclear inner membrane / retrograde protein transport, ER to cytosol / ERAD pathway / RING-type E3 ubiquitin transferase / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / nuclear envelope / protein ubiquitination / endoplasmic reticulum membrane ...Doa10p ubiquitin ligase complex / nuclear inner membrane / retrograde protein transport, ER to cytosol / ERAD pathway / RING-type E3 ubiquitin transferase / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / nuclear envelope / protein ubiquitination / endoplasmic reticulum membrane / endoplasmic reticulum / zinc ion binding / membrane
Similarity search - Function
RING-variant domain / Zinc finger RING-CH-type profile. / Zinc finger, RING-CH-type / The RING-variant domain is a C4HC3 zinc-finger like motif found in a number of cellular and viral proteins. Some of these proteins have been shown both in vivo and in vitro to have ubiquitin E3 ligase activity. / Zinc finger, RING/FYVE/PHD-type
Similarity search - Domain/homology
1,2-DIOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE / ERAD-associated E3 ubiquitin-protein ligase DOA10
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.58 Å
AuthorsBotsch, J.J. / Braeuning, B. / Schulman, B.A.
Funding support Germany, 1items
OrganizationGrant numberCountry
Max Planck Society Germany
CitationJournal: Nat Commun / Year: 2024
Title: Doa10/MARCH6 architecture interconnects E3 ligase activity with lipid-binding transmembrane channel to regulate SQLE.
Authors: J Josephine Botsch / Roswitha Junker / Michèle Sorgenfrei / Patricia P Ogger / Luca Stier / Susanne von Gronau / Peter J Murray / Markus A Seeger / Brenda A Schulman / Bastian Bräuning /
Abstract: Transmembrane E3 ligases play crucial roles in homeostasis. Much protein and organelle quality control, and metabolic regulation, are determined by ER-resident MARCH6 E3 ligases, including Doa10 in ...Transmembrane E3 ligases play crucial roles in homeostasis. Much protein and organelle quality control, and metabolic regulation, are determined by ER-resident MARCH6 E3 ligases, including Doa10 in yeast. Here, we present Doa10/MARCH6 structural analysis by cryo-EM and AlphaFold predictions, and a structure-based mutagenesis campaign. The majority of Doa10/MARCH6 adopts a unique circular structure within the membrane. This channel is established by a lipid-binding scaffold, and gated by a flexible helical bundle. The ubiquitylation active site is positioned over the channel by connections between the cytosolic E3 ligase RING domain and the membrane-spanning scaffold and gate. Here, by assaying 95 MARCH6 variants for effects on stability of the well-characterized substrate SQLE, which regulates cholesterol levels, we reveal crucial roles of the gated channel and RING domain consistent with AlphaFold-models of substrate-engaged and ubiquitylation complexes. SQLE degradation further depends on connections between the channel and RING domain, and lipid binding sites, revealing how interconnected Doa10/MARCH6 elements could orchestrate metabolic signals, substrate binding, and E3 ligase activity.
History
DepositionJun 11, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jan 17, 2024Provider: repository / Type: Initial release

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: ERAD-associated E3 ubiquitin-protein ligase DOA10
hetero molecules


Theoretical massNumber of molelcules
Total (without water)154,6175
Polymers151,6081
Non-polymers3,0094
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area5360 Å2
ΔGint-41 kcal/mol
Surface area53010 Å2

-
Components

#1: Protein ERAD-associated E3 ubiquitin-protein ligase DOA10 / RING-type E3 ubiquitin transferase DOA10


Mass: 151608.109 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: SSM4, DOA10, YIL030C, YI3299.01C, YI9905.18C / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P40318, RING-type E3 ubiquitin transferase
#2: Chemical ChemComp-PCW / 1,2-DIOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / (Z,Z)-4-HYDROXY-N,N,N-TRIMETHYL-10-OXO-7-[(1-OXO-9-OCTADECENYL)OXY]-3,5,9-TRIOXA-4-PHOSPHAHEPTACOS-18-EN-1-AMINIUM-4-OXIDE


Mass: 787.121 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C44H85NO8P / Comment: DOPC, phospholipid*YM
#3: Chemical ChemComp-PX6 / 1,2-DIPALMITOYL-SN-GLYCERO-3-PHOSPHATE


Mass: 647.883 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C35H68O8P
Has ligand of interestN

-
Experimental details

-
Experiment

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

-
Sample preparation

ComponentName: Doa10 with Ubc6 and sybody in MSP1E3D1 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.15 MDa / Experimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

-
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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2800 nm / Nominal defocus min: 700 nm
Image recordingElectron dose: 70 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

-
Processing

EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.58 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 123143 / Symmetry type: POINT
Atomic model buildingSource name: AlphaFold / Type: in silico model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0026407
ELECTRON MICROSCOPYf_angle_d0.4728717
ELECTRON MICROSCOPYf_dihedral_angle_d5.0781065
ELECTRON MICROSCOPYf_chiral_restr0.0371049
ELECTRON MICROSCOPYf_plane_restr0.0041077

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
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.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
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.
  • 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.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
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.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

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

Read more