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- PDB-9mqe: Vitamin K-dependent gamma-carboxylase with Osteocalcin and vitami... -

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Basic information

Entry
Database: PDB / ID: 9mqe
TitleVitamin K-dependent gamma-carboxylase with Osteocalcin and vitamin K hydroquinone
Components
  • Osteocalcin
  • Vitamin K-dependent gamma-carboxylase
KeywordsTRANSFERASE / GGCX / VKGC / Vitamin K / VKCFD / Hemophilia B / Warfarin / Carboxylation / Blood Coagulation / Calcium homeostasis / Osteocalcin / MEMBRANE PROTEIN
Function / homology
Function and homology information


response to hydroxyisoflavone / structural constituent of bone / hydroxyapatite binding / peptidyl-glutamate 4-carboxylase / gamma-glutamyl carboxylase activity / negative regulation of testosterone biosynthetic process / negative regulation of bone development / cellular response to zinc ion starvation / response to macrophage colony-stimulating factor / Defective gamma-carboxylation of F9 ...response to hydroxyisoflavone / structural constituent of bone / hydroxyapatite binding / peptidyl-glutamate 4-carboxylase / gamma-glutamyl carboxylase activity / negative regulation of testosterone biosynthetic process / negative regulation of bone development / cellular response to zinc ion starvation / response to macrophage colony-stimulating factor / Defective gamma-carboxylation of F9 / vitamin binding / vitamin K metabolic process / regulation of testosterone biosynthetic process / negative regulation of neurotransmitter secretion / response to vitamin K / regulation of osteoclast differentiation / cellular response to vitamin D / regulation of bone mineralization / type B pancreatic cell proliferation / regulation of bone resorption / osteoblast development / response to vitamin D / response to zinc ion / positive regulation of neurotransmitter secretion / response to gravity / response to testosterone / bone mineralization / RUNX2 regulates osteoblast differentiation / response to mechanical stimulus / Transport of gamma-carboxylated protein precursors from the endoplasmic reticulum to the Golgi apparatus / Gamma-carboxylation of protein precursors / Removal of aminoterminal propeptides from gamma-carboxylated proteins / regulation of cellular response to insulin stimulus / response to glucocorticoid / protein maturation / response to activity / skeletal system development / stem cell differentiation / hormone activity / bone development / brain development / cellular response to growth factor stimulus / protein modification process / Golgi lumen / response to estrogen / cognition / cellular response to insulin stimulus / blood coagulation / osteoblast differentiation / glucose homeostasis / response to ethanol / perikaryon / vesicle / learning or memory / cell adhesion / response to xenobiotic stimulus / endoplasmic reticulum lumen / calcium ion binding / dendrite / endoplasmic reticulum membrane / structural molecule activity / extracellular space / extracellular region / membrane / cytoplasm
Similarity search - Function
Osteocalcin/matrix Gla protein / Osteocalcin / Vitamin K-dependent gamma-carboxylase / HTTM / : / : / HTTM domain / Vitamin K-dependent gamma-carboxylase, lumenal domain / Horizontally Transferred TransMembrane Domain / RmlC-like cupin domain superfamily ...Osteocalcin/matrix Gla protein / Osteocalcin / Vitamin K-dependent gamma-carboxylase / HTTM / : / : / HTTM domain / Vitamin K-dependent gamma-carboxylase, lumenal domain / Horizontally Transferred TransMembrane Domain / RmlC-like cupin domain superfamily / Gamma-carboxyglutamic acid-rich (GLA) domain / Gamma-carboxyglutamic acid-rich (GLA) domain superfamily / Vitamin K-dependent carboxylation domain. / Gla domain profile. / Domain containing Gla (gamma-carboxyglutamate) residues. / RmlC-like jelly roll fold
Similarity search - Domain/homology
Chem-6PL / : / Osteocalcin / Vitamin K-dependent gamma-carboxylase
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.56 Å
AuthorsLi, W. / Liu, B. / Cao, Q.
Funding support United States, 7items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL121718 United States
American Heart AssociationEstablished Investigator Award United States
American Heart AssociationCollaborative Sciences Award United States
W. M. Keck FoundationForefront of Science Award United States
Childrens Discovery Institute of Washington University and St. Louis Childrens HospitalMCII 2020-854 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI158500 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM131008 United States
CitationJournal: Cell Res / Year: 2025
Title: Structural insights into the vitamin K-dependent γ-carboxylation of osteocalcin.
Authors: Qing Cao / Jianjun Fan / Aaron Ammerman / Samjhana Awasthi / Zongtao Lin / Saimi Mierxiati / Huaping Chen / Jinbin Xu / Benjamin A Garcia / Bin Liu / Weikai Li /
Abstract: The γ-carboxylation state of osteocalcin determines its essential functions in bone mineralization or systemic metabolism and serves as a prominent biomarker for bone health and vitamin K nutrition. ...The γ-carboxylation state of osteocalcin determines its essential functions in bone mineralization or systemic metabolism and serves as a prominent biomarker for bone health and vitamin K nutrition. This post-translational modification of glutamate residues is catalyzed by the membrane-embedded vitamin K-dependent γ-carboxylase (VKGC), which typically recognizes protein substrates through their tightly bound propeptide that triggers γ-carboxylation. However, the osteocalcin propeptide exhibits negligible affinity for VKGC. To understand the underlying molecular mechanism, we determined the cryo-electron microscopy structures of VKGC with osteocalcin carrying a native propeptide or a high-affinity variant at different carboxylation states. The structures reveal a large chamber in VKGC that maintains uncarboxylated and partially carboxylated osteocalcin in partially unfolded conformations, allowing their glutamate-rich region and C-terminal helices to engage with VKGC at multiple sites. Binding of this mature region together with the low-affinity propeptide effectively stimulates VKGC activity, similar to high-affinity propeptides that differ only in closely fitting interactions. However, the low-affinity propeptide renders osteocalcin prone to undercarboxylation at low vitamin K levels, thereby serving as a discernible biomarker. Overall, our studies reveal the unique interaction of osteocalcin with VKGC and provide a framework for designing therapeutic strategies targeting osteocalcin-related bone and metabolic disorders.
History
DepositionJan 2, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 8, 2025Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Vitamin K-dependent gamma-carboxylase
P: Osteocalcin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)102,12310
Polymers98,6312
Non-polymers3,4918
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 2 types, 2 molecules AP

#1: Protein Vitamin K-dependent gamma-carboxylase / Gamma-glutamyl carboxylase / Peptidyl-glutamate 4-carboxylase / Vitamin K gamma glutamyl carboxylase


Mass: 87655.641 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GGCX, GC / Cell line (production host): HEK293S / Production host: Homo sapiens (human)
References: UniProt: P38435, peptidyl-glutamate 4-carboxylase
#2: Protein Osteocalcin / Bone Gla protein / BGP / Gamma-carboxyglutamic acid-containing protein


Mass: 10975.558 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: BGLAP / Cell line (production host): HEK293S / Production host: Homo sapiens (human) / References: UniProt: P02818

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Sugars , 2 types, 5 molecules

#3: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#4: Sugar ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 2 types, 3 molecules

#5: Chemical ChemComp-6PL / (4S,7R)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE / 1-PALMITOYL-2-STEAROYL-SN-GLYCERO-3-PHOSPHOCHOLINE


Mass: 763.100 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C42H85NO8P / Feature type: SUBJECT OF INVESTIGATION / Comment: phospholipid*YM
#6: Chemical ChemComp-A1AVC / vitamin K1 hydroquinone / 2-methyl-3-[(2E,7R,11R)-3,7,11,15-tetramethylhexadec-2-en-1-yl]naphthalene-1,4-diol


Mass: 452.712 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C31H48O2 / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
Has protein modificationY

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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: Vitamin K-dependent gamma-carboxylase with Osteocalcin and vitamin K hydroquinone
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 0.11 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293S / Plasmid: pEG BacMam
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMSodium chlorideNaCl1
220 mMHEPESC8H18N2O4S1
30.005 %Glyco-diosgeninC56H92O251
40.055 mMVitamin K hydroquinoneC31H48O21
51.84 mMSodium bicarbonateNaHCO31
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 283.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 77 K / Temperature (min): 63 K
Image recordingElectron dose: 47 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 7089
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansWidth: 4092 / Height: 5760

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2PHENIX1.21.1_5286:model refinement
3EPUimage acquisition
5cryoSPARCCTF correction
10cryoSPARCinitial Euler assignment
11cryoSPARCfinal Euler assignment
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1322129
3D reconstructionResolution: 3.56 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 150446 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0046580
ELECTRON MICROSCOPYf_angle_d0.7368910
ELECTRON MICROSCOPYf_dihedral_angle_d11.5141082
ELECTRON MICROSCOPYf_chiral_restr0.041959
ELECTRON MICROSCOPYf_plane_restr0.0051110

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