[English] 日本語
Yorodumi
- EMDB-48594: Yeast V-ATPase bound to Nanobody 2WVA7, state 3+18 -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-48594
TitleYeast V-ATPase bound to Nanobody 2WVA7, state 3+18
Map dataYeast_V1Vo_N27_state3 18_main
Sample
  • Complex: Yeast V-ATPase Vo proton channel bound to nanobody 2WVA7
KeywordsVacuolar ATPase / Vo proton channel / lipid nanodisc / nanobody / PROTON TRANSPORT
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsWilkens S / Knight K / Roh S-H / Park JB
Funding support United States, Korea, Republic Of, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM141908 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM058600 United States
National Research Foundation (NRF, Korea)2020R1A6C101A183 Korea, Republic Of
National Research Foundation (NRF, Korea)2021M3A9I4021220 Korea, Republic Of
CitationJournal: bioRxiv / Year: 2025
Title: Monoclonal nanobodies alter the activity and assembly of the yeast vacuolar H-ATPase.
Authors: Kassidy Knight / Jun Bae Park / Rebecca A Oot / Md Murad Khan / Soung-Hun Roh / Stephan Wilkens /
Abstract: The vacuolar ATPase (V-ATPase; VV) is a multi-subunit rotary nanomotor proton pump that acidifies organelles in virtually all eukaryotic cells, and extracellular spaces in some specialized tissues of ...The vacuolar ATPase (V-ATPase; VV) is a multi-subunit rotary nanomotor proton pump that acidifies organelles in virtually all eukaryotic cells, and extracellular spaces in some specialized tissues of higher organisms. Evidence suggests that metastatic breast cancers mislocalize V-ATPase to the plasma membrane to promote cell survival and facilitate metastasis, making the V-ATPase a potential drug target. We have generated a library of camelid single-domain antibodies (Nanobodies; Nbs) against lipid-nanodisc reconstituted yeast V-ATPase V proton channel subcomplex. Here, we present an in-depth characterization of three anti-V Nbs using biochemical and biophysical experiments. We find that the Nbs bind V with high affinity, with one Nb inhibiting holoenzyme activity and another one preventing enzyme assembly. Using cryoEM, we find that two of the Nbs bind the subunit ring of the V on the lumen side of the complex. Additionally, we show that one of the Nbs raised against yeast V can pull down human V-ATPase (VV). Our research demonstrates Nb versatility to target and modulate the activity of the V-ATPase, and highlights the potential for future therapeutic Nb development.
History
DepositionJan 10, 2025-
Header (metadata) releaseFeb 5, 2025-
Map releaseFeb 5, 2025-
UpdateFeb 5, 2025-
Current statusFeb 5, 2025Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_48594.png

Downloads & links

-
Map

FileDownload / File: emd_48594.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationYeast_V1Vo_N27_state3 18_main
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 320 pix.
= 352. Å		1.1 Å/pix.
x 320 pix.
= 352. Å		1.1 Å/pix.
x 320 pix.
= 352. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.004
Minimum - Maximum-0.010839089 - 0.025934845
Average (Standard dev.)0.000022817847 (±0.0014382045)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 352.0 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Mask #1

Fileemd_48594_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Yeast V1Vo N27 state3 18 half1

Fileemd_48594_half_map_1.map
AnnotationYeast_V1Vo_N27_state3 18_half1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Yeast V1Vo N27 state3 18 half2

Fileemd_48594_half_map_2.map
AnnotationYeast_V1Vo_N27_state3 18_half2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Yeast V-ATPase Vo proton channel bound to nanobody 2WVA7

EntireName: Yeast V-ATPase Vo proton channel bound to nanobody 2WVA7
Components
  • Complex: Yeast V-ATPase Vo proton channel bound to nanobody 2WVA7

-
Supramolecule #1: Yeast V-ATPase Vo proton channel bound to nanobody 2WVA7

SupramoleculeName: Yeast V-ATPase Vo proton channel bound to nanobody 2WVA7
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#9
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 540 KDa

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration2 mg/mL
BufferpH: 7.4
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER

-
Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Average electron dose: 39.96 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.3 µm

+
Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 5) / Number images used: 30492
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 5)
FSC plot (resolution estimation)

+
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