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- PDB-7zbn: Cryo-EM structure of the human GS-GN complex in the inhibited state -

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Basic information

Entry
Database: PDB / ID: 7zbn
TitleCryo-EM structure of the human GS-GN complex in the inhibited state
Components
  • (Glycogen [starch] synthase, muscle) x 2
  • Isoform GN-1 of Glycogenin-1
KeywordsTRANSFERASE / Glycosyltransferase
Function / homology
Function and homology information


glycogen synthase activity, transferring glucose-1-phosphate / Glycogen storage disease type XV (GYG1) / Glycogen storage disease type 0 (muscle GYS1) / glycogen(starch) synthase / Glycogen storage disease type II (GAA) / glycogenin glucosyltransferase / glycogenin glucosyltransferase activity / UDP-alpha-D-glucose:glucosyl-glycogenin alpha-D-glucosyltransferase activity / glycogen (starch) synthase activity / D-glucose binding ...glycogen synthase activity, transferring glucose-1-phosphate / Glycogen storage disease type XV (GYG1) / Glycogen storage disease type 0 (muscle GYS1) / glycogen(starch) synthase / Glycogen storage disease type II (GAA) / glycogenin glucosyltransferase / glycogenin glucosyltransferase activity / UDP-alpha-D-glucose:glucosyl-glycogenin alpha-D-glucosyltransferase activity / glycogen (starch) synthase activity / D-glucose binding / glycogen biosynthetic process / Glycogen breakdown (glycogenolysis) / glycosyltransferase activity / inclusion body / Myoclonic epilepsy of Lafora / Glycogen synthesis / lysosomal lumen / manganese ion binding / heart development / secretory granule lumen / ficolin-1-rich granule lumen / Neutrophil degranulation / protein homodimerization activity / extracellular region / membrane / cytosol / cytoplasm
Similarity search - Function
Glycogen synthase / Glycogen synthase / Glycosyl transferase, family 8 / Glycosyl transferase family 8 / Nucleotide-diphospho-sugar transferases
Similarity search - Domain/homology
Glycogen [starch] synthase, muscle / Glycogenin-1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.62 Å
AuthorsMarr, L. / Zeqiraj, E.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
UK Research and Innovation (UKRI) United Kingdom
CitationJournal: Nat Commun / Year: 2022
Title: Mechanism of glycogen synthase inactivation and interaction with glycogenin.
Authors: Laura Marr / Dipsikha Biswas / Leonard A Daly / Christopher Browning / Sarah C M Vial / Daniel P Maskell / Catherine Hudson / Jay A Bertrand / John Pollard / Neil A Ranson / Heena Khatter / ...Authors: Laura Marr / Dipsikha Biswas / Leonard A Daly / Christopher Browning / Sarah C M Vial / Daniel P Maskell / Catherine Hudson / Jay A Bertrand / John Pollard / Neil A Ranson / Heena Khatter / Claire E Eyers / Kei Sakamoto / Elton Zeqiraj /
Abstract: Glycogen is the major glucose reserve in eukaryotes, and defects in glycogen metabolism and structure lead to disease. Glycogenesis involves interaction of glycogenin (GN) with glycogen synthase (GS) ...Glycogen is the major glucose reserve in eukaryotes, and defects in glycogen metabolism and structure lead to disease. Glycogenesis involves interaction of glycogenin (GN) with glycogen synthase (GS), where GS is activated by glucose-6-phosphate (G6P) and inactivated by phosphorylation. We describe the 2.6 Å resolution cryo-EM structure of phosphorylated human GS revealing an autoinhibited GS tetramer flanked by two GN dimers. Phosphorylated N- and C-termini from two GS protomers converge near the G6P-binding pocket and buttress against GS regulatory helices. This keeps GS in an inactive conformation mediated by phospho-Ser641 interactions with a composite "arginine cradle". Structure-guided mutagenesis perturbing interactions with phosphorylated tails led to increased basal/unstimulated GS activity. We propose that multivalent phosphorylation supports GS autoinhibition through interactions from a dynamic "spike" region, allowing a tuneable rheostat for regulating GS activity. This work therefore provides insights into glycogen synthesis regulation and facilitates studies of glycogen-related diseases.
History
DepositionMar 23, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 22, 2022Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Glycogen [starch] synthase, muscle
B: Glycogen [starch] synthase, muscle
E: Isoform GN-1 of Glycogenin-1
F: Isoform GN-1 of Glycogenin-1
C: Glycogen [starch] synthase, muscle
D: Glycogen [starch] synthase, muscle
G: Isoform GN-1 of Glycogenin-1
H: Isoform GN-1 of Glycogenin-1


Theoretical massNumber of molelcules
Total (without water)485,5798
Polymers485,5798
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: equilibrium centrifugation
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area21270 Å2
ΔGint-108 kcal/mol
Surface area91620 Å2

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Components

#1: Protein Glycogen [starch] synthase, muscle


Mass: 83885.430 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GYS1, GYS / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P13807, glycogen(starch) synthase
#2: Protein Glycogen [starch] synthase, muscle


Mass: 83965.406 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GYS1, GYS / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P13807, glycogen(starch) synthase
#3: Protein
Isoform GN-1 of Glycogenin-1 / GN-1 / GN1


Mass: 37469.348 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GYG1, GYG / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P46976, glycogenin glucosyltransferase
Has ligand of interestY

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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: Glycogen synthase-Glycogenin complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.485 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMSodium ChlorideNaClSodium chloride1
225 mMHEPES1
31 mMTCEP1
41.5 %Glycerol1
SpecimenConc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2200 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 34.8 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM softwareName: RELION / Category: 3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.62 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 739232 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00721672
ELECTRON MICROSCOPYf_angle_d0.61829356
ELECTRON MICROSCOPYf_dihedral_angle_d5.2692886
ELECTRON MICROSCOPYf_chiral_restr0.0453110
ELECTRON MICROSCOPYf_plane_restr0.0053790

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