[English] 日本語
Yorodumi
- EMDB-24784: Cryo-EM map of human GlcNAc-1-phosphotransferase A2B2 subcomplex -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-24784
TitleCryo-EM map of human GlcNAc-1-phosphotransferase A2B2 subcomplex
Map data
Sample
  • Complex: GlcNAc-1-phosphotransferase
    • Protein or peptide: N-acetylglucosamine-1-phosphotransferase subunits alpha/beta
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: CALCIUM ION
KeywordsGlcNAc-1-phosphotransferase / lysosomal hydrolases / mannose 6-phosphate trafficking pathway / TRANSFERASE
Function / homology
Function and homology information


UDP-N-acetylglucosamine-lysosomal-enzyme N-acetylglucosaminephosphotransferase / N-glycan processing to lysosome / UDP-N-acetylglucosamine-lysosomal-enzyme N-acetylglucosaminephosphotransferase activity / secretion of lysosomal enzymes / carbohydrate phosphorylation / lysosome organization / Golgi membrane / calcium ion binding / Golgi apparatus
Similarity search - Function
N-acetylglucosamine-1-phosphotransferase subunit alpha/beta, regulatory domain / Putative GlcNAc-1 phosphotransferase regulatory domain / Stealth protein CR2, conserved region 2 / Stealth protein CR4, conserved region 4 / Stealth protein CR3, conserved region 3 / Stealth protein CR1, conserved region 1 / : / Stealth protein CR2, conserved region 2 / Stealth protein CR1, conserved region 1 / Stealth protein CR3, conserved region 3 ...N-acetylglucosamine-1-phosphotransferase subunit alpha/beta, regulatory domain / Putative GlcNAc-1 phosphotransferase regulatory domain / Stealth protein CR2, conserved region 2 / Stealth protein CR4, conserved region 4 / Stealth protein CR3, conserved region 3 / Stealth protein CR1, conserved region 1 / : / Stealth protein CR2, conserved region 2 / Stealth protein CR1, conserved region 1 / Stealth protein CR3, conserved region 3 / Stealth protein CR4, conserved region 4 / DMAP1-binding Domain / DMAP1-binding Domain / DMAP1-binding domain / DMAP1-binding domain profile. / Notch-like domain superfamily / LNR domain / LNR (Lin-12/Notch) repeat profile. / Notch domain / Domain found in Notch and Lin-12 / EF-Hand 1, calcium-binding site / EF-hand calcium-binding domain. / EF-hand calcium-binding domain profile. / EF-hand domain
Similarity search - Domain/homology
N-acetylglucosamine-1-phosphotransferase subunits alpha/beta
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsLi H
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)CA231466 United States
CitationJournal: Nat Struct Mol Biol / Year: 2022
Title: Structure of the human GlcNAc-1-phosphotransferase αβ subunits reveals regulatory mechanism for lysosomal enzyme glycan phosphorylation.
Authors: Hua Li / Wang-Sik Lee / Xiang Feng / Lin Bai / Benjamin C Jennings / Lin Liu / Balraj Doray / William M Canfield / Stuart Kornfeld / Huilin Li /
Abstract: Vertebrates use the mannose 6-phosphate (M6P)-recognition system to deliver lysosomal hydrolases to lysosomes. Key to this pathway is N-acetylglucosamine (GlcNAc)-1-phosphotransferase (PTase) that ...Vertebrates use the mannose 6-phosphate (M6P)-recognition system to deliver lysosomal hydrolases to lysosomes. Key to this pathway is N-acetylglucosamine (GlcNAc)-1-phosphotransferase (PTase) that selectively adds GlcNAc-phosphate (P) to mannose residues of hydrolases. Human PTase is an αβγ heterohexamer with a catalytic core and several peripheral domains that recognize and bind substrates. Here we report a cryo-EM structure of the catalytic core of human PTase and the identification of a hockey stick-like motif that controls activation of the enzyme. Movement of this motif out of the catalytic pocket is associated with a rearrangement of part of the peripheral domains that unblocks hydrolase glycan access to the catalytic site, thereby activating PTase. We propose that PTase fluctuates between inactive and active states in solution, and selective substrate binding of a lysosomal hydrolase through its protein-binding determinant to PTase locks the enzyme in the active state to permit glycan phosphorylation. This mechanism would help ensure that only N-linked glycans of lysosomal enzymes are phosphorylated.
History
DepositionAug 30, 2021-
Header (metadata) releaseMar 30, 2022-
Map releaseMar 30, 2022-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_24784.png

Downloads & links

-
Map

FileDownload / File: emd_24784.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 320 pix.
= 264.96 Å		0.83 Å/pix.
x 320 pix.
= 264.96 Å		0.83 Å/pix.
x 320 pix.
= 264.96 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.828 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-3.3548815 - 4.380209
Average (Standard dev.)0.0036207975 (±0.07669943)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 264.96 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: #2

Fileemd_24784_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: #1

Fileemd_24784_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : GlcNAc-1-phosphotransferase

EntireName: GlcNAc-1-phosphotransferase
Components
  • Complex: GlcNAc-1-phosphotransferase
    • Protein or peptide: N-acetylglucosamine-1-phosphotransferase subunits alpha/beta
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: CALCIUM ION

-
Supramolecule #1: GlcNAc-1-phosphotransferase

SupramoleculeName: GlcNAc-1-phosphotransferase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

-
Macromolecule #1: N-acetylglucosamine-1-phosphotransferase subunits alpha/beta

MacromoleculeName: N-acetylglucosamine-1-phosphotransferase subunits alpha/beta
type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
EC number: UDP-N-acetylglucosamine-lysosomal-enzyme N-acetylglucosaminephosphotransferase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 134.7875 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: DEDQVDPRLI DGKWSRDQYH VLFDSYRDNI AGKSFQNRLC LPMPIDVVYT WVNGTDLELL KELQQVREQM EEEQKAMREI LGKNTTEPT KKSEKQLECL LTHCIKVPML VLDPALPANI TLKDLPSLYP SFHSASDIFN VAKPKNPSTN VSVVVFDSTK D VEDAHSGL ...String:
DEDQVDPRLI DGKWSRDQYH VLFDSYRDNI AGKSFQNRLC LPMPIDVVYT WVNGTDLELL KELQQVREQM EEEQKAMREI LGKNTTEPT KKSEKQLECL LTHCIKVPML VLDPALPANI TLKDLPSLYP SFHSASDIFN VAKPKNPSTN VSVVVFDSTK D VEDAHSGL LKGNSRQTVW RGYLTTDKEV PGLVLMQDLA FLSGFPPTFK ETNQLKTKLP ENLSSKVKLL QLYSEASVAL LK LNNPKDF QELNKQTKKN MTIDGKELTI SPAYLLWDLS AISQSKQDED ISASRFEDNE ELRYSLRSIE RHAPWVRNIF IVT NGQIPS WLNLDNPRVT IVTHQDVFRN LSHLPTFSSP AIESHIHRIE GLSQKFIYLN DDVMFGKDVW PDDFYSHSKG QKVY LTWPV PNCAEGCPGS WIKDGYCDKA CNNSACDWDG GDCSGNSGGS RYIAGGGGTG SIGVGQPWQF GGGINSVSYC NQGCA NSWL ADKFCDQACN VLSCGFDAGD CGQDHFHELY KVILLPNQTH YIIPKGECLP YFSFAEVAKR GVEGAYSDNP IIRHAS IAN KWKTIHLIMH SGMNATTIHF NLTFQNTNDE EFKMQITVEV DTREGPKLNS TAQKGYENLV SPITLLPEAE ILFEDIP KE KRFPKFKRHD VNSTRRAQEE VKIPLVNISL LPKDAQLSLN TLDLQLEHGD ITLKGYNLSK SALLRSFLMN SQHAKIKN Q AIITDETNDS LVAPQEKQVH KSILPNSLGV SERLQRLTFP AVSVKVNGHD QGQNPPLDLE TTARFRVETH TQKTIGGNV TKEKPPSLIV PLESQMTKEK KITGKEKENS RMEENAENHI GVTEVLLGRK LQHYTDSYLG FLPWEKKKYF QDLLDEEESL KTQLAYFTD SKNRARYKRD TFADSLRYVN KILNSKFGFT SRKVPAHMPH MIDRIVMQEL QDMFPEEFDK TSFHKVRHSE D MQFAFSYF YYLMSAVQPL NISQVFDEVD TDQSGVLSDR EIRTLATRIH ELPLSLQDLT GLEHMLINCS KMLPADITQL NN IPPTQES YYDPNLPPVT KSLVTNCKPV TDKIHKAYKD KNKYRFEIMG EEEIAFKMIR TNVSHVVGQL DDIRKNPRKF VCL NDNIDH NHKDAQTVKA VLRDFYESMF PIPSQFELPR EYRNRFLHMH ELQEWRAYRD KLK

UniProtKB: N-acetylglucosamine-1-phosphotransferase subunits alpha/beta

-
Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 6 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

-
Macromolecule #4: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: CA
Molecular weightTheoretical: 40.078 Da

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration0.15 mg/mL
BufferpH: 7.8
Component:
ConcentrationFormulaName
75.0 mMKClpotassium chloride
20.0 mMHEPES
10.0 mM2-mercaptoethanol
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 299 K / Instrument: FEI VITROBOT MARK IV

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
TemperatureMin: 193.0 K / Max: 193.0 K
Alignment procedureComa free - Residual tilt: 0.05 mrad
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 13320 / Average exposure time: 1.5 sec. / Average electron dose: 66.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

+
Image processing

Particle selectionNumber selected: 5186047
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.1) / Number images used: 365927
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
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