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- PDB-8jfl: PhK holoenzyme in active state, muscle isoform -

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Basic information

Entry
Database: PDB / ID: 8jfl
TitlePhK holoenzyme in active state, muscle isoform
Components
  • Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform
  • Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform
  • Phosphorylase b kinase regulatory subunit beta
KeywordsCYTOSOLIC PROTEIN / glycogen phosphorylase b kinase / muscle isoform / Ca2+ active state
Function / homology
Function and homology information


phosphorylase kinase / phosphorylase kinase activity / phosphorylase kinase complex / positive regulation of glycogen catabolic process / tau-protein kinase / Glycogen breakdown (glycogenolysis) / glycogen metabolic process / generation of precursor metabolites and energy / carbohydrate metabolic process / calmodulin binding ...phosphorylase kinase / phosphorylase kinase activity / phosphorylase kinase complex / positive regulation of glycogen catabolic process / tau-protein kinase / Glycogen breakdown (glycogenolysis) / glycogen metabolic process / generation of precursor metabolites and energy / carbohydrate metabolic process / calmodulin binding / non-specific serine/threonine protein kinase / protein serine kinase activity / enzyme binding / signal transduction / ATP binding / plasma membrane / cytoplasm / cytosol
Similarity search - Function
Phosphorylase kinase alpha/beta subunit / Phosphorylase b kinase regulatory subunit alpha/beta, C-terminal domain / Phosphorylase b kinase C-terminal domain / Phosphorylase kinase, gamma catalytic subunit / GH15-like domain / Glycosyl hydrolases family 15 / Six-hairpin glycosidase-like superfamily / Six-hairpin glycosidase superfamily / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. ...Phosphorylase kinase alpha/beta subunit / Phosphorylase b kinase regulatory subunit alpha/beta, C-terminal domain / Phosphorylase b kinase C-terminal domain / Phosphorylase kinase, gamma catalytic subunit / GH15-like domain / Glycosyl hydrolases family 15 / Six-hairpin glycosidase-like superfamily / Six-hairpin glycosidase superfamily / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / FARNESYL / Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform / Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform / Phosphorylase b kinase regulatory subunit beta
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsYang, X.K. / Xiao, J.Y.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Nat Commun / Year: 2024
Title: Architecture and activation of human muscle phosphorylase kinase.
Authors: Xiaoke Yang / Mingqi Zhu / Xue Lu / Yuxin Wang / Junyu Xiao /
Abstract: The study of phosphorylase kinase (PhK)-regulated glycogen metabolism has contributed to the fundamental understanding of protein phosphorylation; however, the molecular mechanism of PhK remains ...The study of phosphorylase kinase (PhK)-regulated glycogen metabolism has contributed to the fundamental understanding of protein phosphorylation; however, the molecular mechanism of PhK remains poorly understood. Here we present the high-resolution cryo-electron microscopy structures of human muscle PhK. The 1.3-megadalton PhK αβγδ hexadecamer consists of a tetramer of tetramer, wherein four αβγδ modules are connected by the central β scaffold. The α- and β-subunits possess glucoamylase-like domains, but exhibit no detectable enzyme activities. The α-subunit serves as a bridge between the β-subunit and the γδ subcomplex, and facilitates the γ-subunit to adopt an autoinhibited state. Ca-free calmodulin (δ-subunit) binds to the γ-subunit in a compact conformation. Upon binding of Ca, a conformational change occurs, allowing for the de-inhibition of the γ-subunit through a spring-loaded mechanism. We also reveal an ADP-binding pocket in the β-subunit, which plays a role in allosterically enhancing PhK activity. These results provide molecular insights of this important kinase complex.
History
DepositionMay 18, 2023Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Apr 3, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 10, 2024Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform
C: Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform
E: Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform
G: Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform
M: Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform
O: Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform
I: Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform
K: Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform
B: Phosphorylase b kinase regulatory subunit beta
F: Phosphorylase b kinase regulatory subunit beta
J: Phosphorylase b kinase regulatory subunit beta
N: Phosphorylase b kinase regulatory subunit beta
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,233,70824
Polymers1,230,34812
Non-polymers3,36012
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Phosphorylase b kinase regulatory subunit alpha, skeletal muscle isoform / Phosphorylase kinase alpha M subunit


Mass: 137469.422 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PHKA1, PHKA / Production host: Homo sapiens (human) / References: UniProt: P46020
#2: Protein
Phosphorylase b kinase gamma catalytic chain, skeletal muscle/heart isoform / PHK-gamma-M / Phosphorylase kinase subunit gamma-1 / Serine/threonine-protein kinase PHKG1


Mass: 45084.672 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PHKG1, PHKG / Production host: Homo sapiens (human)
References: UniProt: Q16816, phosphorylase kinase, non-specific serine/threonine protein kinase, tau-protein kinase
#3: Protein
Phosphorylase b kinase regulatory subunit beta / Phosphorylase kinase subunit beta


Mass: 125032.961 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PHKB / Production host: Homo sapiens (human) / References: UniProt: Q93100
#4: Chemical
ChemComp-FAR / FARNESYL


Mass: 206.367 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: C15H26 / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
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: phosphorylase b kinase, muscle isoform, Ca2+ active state
Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8.2
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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: SPOT SCAN
Electron lensMode: OTHER / Nominal defocus max: 1500 nm / Nominal defocus min: 1100 nm
Image recordingElectron dose: 1.5 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 432047 / Symmetry type: POINT

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