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- PDB-8jut: rat megalin RAP complex -

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Basic information

Entry
Database: PDB / ID: 8jut
Titlerat megalin RAP complex
Components
  • (unclear peptide) x 6
  • Alpha-2-macroglobulin receptor-associated protein
  • LDL receptor related protein 2
KeywordsENDOCYTOSIS / endocytosis receptor
Function / homology
Function and homology information


lipase binding / negative regulation of very-low-density lipoprotein particle clearance / regulation of receptor-mediated endocytosis / rough endoplasmic reticulum lumen / receptor antagonist activity / amyloid-beta clearance by transcytosis / negative regulation of amyloid-beta clearance / hormone binding / very-low-density lipoprotein particle receptor binding / cis-Golgi network ...lipase binding / negative regulation of very-low-density lipoprotein particle clearance / regulation of receptor-mediated endocytosis / rough endoplasmic reticulum lumen / receptor antagonist activity / amyloid-beta clearance by transcytosis / negative regulation of amyloid-beta clearance / hormone binding / very-low-density lipoprotein particle receptor binding / cis-Golgi network / negative regulation of receptor internalization / low-density lipoprotein particle receptor binding / endoplasmic reticulum-Golgi intermediate compartment / endomembrane system / receptor-mediated endocytosis / endosome lumen / Golgi lumen / heparin binding / vesicle / receptor ligand activity / receptor complex / endosome / apical plasma membrane / calcium ion binding / Golgi apparatus / cell surface / signal transduction / plasma membrane
Similarity search - Function
Alpha-2-macroglobulin receptor-associated protein, domain 1 / Alpha-2-macroglobulin RAP, C-terminal / RAP domain superfamily / Alpha-2-macroglobulin RAP, domain 3 / Alpha-2-macroglobulin RAP, domain 2 / Alpha-2-macroglobulin receptor-associated protein / Alpha-2-macroglobulin RAP, N-terminal domain / Alpha-2-macroglobulin RAP, C-terminal domain / Complement Clr-like EGF domain / Complement Clr-like EGF-like ...Alpha-2-macroglobulin receptor-associated protein, domain 1 / Alpha-2-macroglobulin RAP, C-terminal / RAP domain superfamily / Alpha-2-macroglobulin RAP, domain 3 / Alpha-2-macroglobulin RAP, domain 2 / Alpha-2-macroglobulin receptor-associated protein / Alpha-2-macroglobulin RAP, N-terminal domain / Alpha-2-macroglobulin RAP, C-terminal domain / Complement Clr-like EGF domain / Complement Clr-like EGF-like / Low-density lipoprotein receptor repeat class B / LDL-receptor class B (LDLRB) repeat profile. / LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Low-density lipoprotein receptor domain class A / Endoplasmic reticulum targeting sequence. / Low-density lipoprotein (LDL) receptor class A, conserved site / LDL-receptor class A (LDLRA) domain signature. / LDL-receptor class A (LDLRA) domain profile. / : / Calcium-binding EGF domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A repeat / LDL receptor-like superfamily / Six-bladed beta-propeller, TolB-like / Coagulation Factor Xa inhibitory site / EGF-type aspartate/asparagine hydroxylation site / EGF-like calcium-binding, conserved site / Calcium-binding EGF-like domain signature. / Aspartic acid and asparagine hydroxylation site. / EGF-like calcium-binding domain / Calcium-binding EGF-like domain / Epidermal growth factor-like domain. / EGF-like domain profile. / Growth factor receptor cysteine-rich domain superfamily / EGF-like domain signature 2. / EGF-like domain signature 1. / EGF-like domain / Prokaryotic membrane lipoprotein lipid attachment site profile.
Similarity search - Domain/homology
2-acetamido-2-deoxy-alpha-D-galactopyranose / NICKEL (II) ION / LDL receptor related protein 2 / Alpha-2-macroglobulin receptor-associated protein
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsGoto, S. / Tsutsumi, A. / Lee, Y. / Hosojima, M. / Kabasawa, H. / Komochi, K. / Yun-san, L. / Nagatoshi, S. / Tsumoto, K. / Nishizawa, T. ...Goto, S. / Tsutsumi, A. / Lee, Y. / Hosojima, M. / Kabasawa, H. / Komochi, K. / Yun-san, L. / Nagatoshi, S. / Tsumoto, K. / Nishizawa, T. / Kikkawa, M. / Saito, A.
Funding support Japan, 1items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS) Japan
CitationJournal: Proc Natl Acad Sci U S A / Year: 2024
Title: Cryo-EM structures elucidate the multiligand receptor nature of megalin.
Authors: Sawako Goto / Akihisa Tsutsumi / Yongchan Lee / Michihiro Hosojima / Hideyuki Kabasawa / Koichi Komochi / Satoru Nagatoishi / Kazuya Takemoto / Kouhei Tsumoto / Tomohiro Nishizawa / Masahide ...Authors: Sawako Goto / Akihisa Tsutsumi / Yongchan Lee / Michihiro Hosojima / Hideyuki Kabasawa / Koichi Komochi / Satoru Nagatoishi / Kazuya Takemoto / Kouhei Tsumoto / Tomohiro Nishizawa / Masahide Kikkawa / Akihiko Saito /
Abstract: Megalin (low-density lipoprotein receptor-related protein 2) is a giant glycoprotein of about 600 kDa, mediating the endocytosis of more than 60 ligands, including those of proteins, peptides, and ...Megalin (low-density lipoprotein receptor-related protein 2) is a giant glycoprotein of about 600 kDa, mediating the endocytosis of more than 60 ligands, including those of proteins, peptides, and drug compounds [S. Goto, M. Hosojima, H. Kabasawa, A. Saito, , 106393 (2023)]. It is expressed predominantly in renal proximal tubule epithelial cells, as well as in the brain, lungs, eyes, inner ear, thyroid gland, and placenta. Megalin is also known to mediate the endocytosis of toxic compounds, particularly those that cause renal and hearing disorders [Y. Hori , , 1783-1791 (2017)]. Genetic megalin deficiency causes Donnai-Barrow syndrome/facio-oculo-acoustico-renal syndrome in humans. However, it is not known how megalin interacts with such a wide variety of ligands and plays pathological roles in various organs. In this study, we elucidated the dimeric architecture of megalin, purified from rat kidneys, using cryoelectron microscopy. The maps revealed the densities of endogenous ligands bound to various regions throughout the dimer, elucidating the multiligand receptor nature of megalin. We also determined the structure of megalin in complex with receptor-associated protein, a molecular chaperone for megalin. The results will facilitate further studies on the pathophysiology of megalin-dependent multiligand endocytic pathways in multiple organs and will also be useful for the development of megalin-targeted drugs for renal and hearing disorders, Alzheimer's disease [B. V. Zlokovic , , 4229-4234 (1996)], and other illnesses.
History
DepositionJun 27, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 19, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: LDL receptor related protein 2
B: LDL receptor related protein 2
C: Alpha-2-macroglobulin receptor-associated protein
D: Alpha-2-macroglobulin receptor-associated protein
G: unclear peptide
H: unclear peptide
I: unclear peptide
J: unclear peptide
K: unclear peptide
L: unclear peptide
M: unclear peptide
N: unclear peptide
O: unclear peptide
P: unclear peptide
Q: unclear peptide
R: unclear peptide
S: unclear peptide
T: unclear peptide
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,172,518198
Polymers1,130,62018
Non-polymers41,898180
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, 4 molecules ABCD

#1: Protein LDL receptor related protein 2 / rat megalin


Mass: 519871.438 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Rattus norvegicus (Norway rat) / References: UniProt: A0A0G2K9W7
#2: Protein Alpha-2-macroglobulin receptor-associated protein / Alpha-2-MRAP / Gp330-binding 45 kDa protein / Low density lipoprotein receptor-related protein- ...Alpha-2-MRAP / Gp330-binding 45 kDa protein / Low density lipoprotein receptor-related protein-associated protein 1 / RAP


Mass: 42112.809 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Lrpap1 / Production host: Escherichia coli (E. coli) / References: UniProt: Q99068

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Protein/peptide , 6 types, 14 molecules GNHOIPJQKRLMST

#3: Protein/peptide unclear peptide


Mass: 528.644 Da / Num. of mol.: 2 / Source method: isolated from a natural source
Details: Authors state that they could not experimentally confirm this sequence. Therefore residue numbers and chain IDs are meaningless for entity 3-8.
Source: (natural) Rattus norvegicus (Norway rat)
#4: Protein/peptide unclear peptide


Mass: 472.537 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Rattus norvegicus (Norway rat)
#5: Protein/peptide unclear peptide


Mass: 556.697 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Rattus norvegicus (Norway rat)
#6: Protein/peptide unclear peptide


Mass: 291.368 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Rattus norvegicus (Norway rat)
#7: Protein/peptide unclear peptide


Mass: 531.557 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Rattus norvegicus (Norway rat)
#8: Protein/peptide
unclear peptide


Mass: 472.537 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Rattus norvegicus (Norway rat)

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Sugars , 10 types, 90 molecules

#9: 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.: 13 / Source method: obtained synthetically
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
#10: Polysaccharide
alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2- ...alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 910.823 Da / Num. of mol.: 12 / Source method: obtained synthetically
DescriptorTypeProgram
DManpa1-3[DManpa1-6]DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,5,4/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3/a4-b1_b4-c1_c3-d1_c6-e1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}[(6+1)][a-D-Manp]{}}}}LINUCSPDB-CARE
#11: Polysaccharide alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-3)-2-acetamido-2- ...alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 910.823 Da / Num. of mol.: 1 / Source method: obtained synthetically
DescriptorTypeProgram
DManpa1-3[DManpa1-6]DManpb1-3DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,5,4/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3/a4-b1_b3-c1_c3-d1_c6-e1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(3+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}[(6+1)][a-D-Manp]{}}}}LINUCSPDB-CARE
#12: Polysaccharide
beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}LINUCSPDB-CARE
#13: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 3 / Source method: obtained synthetically
DescriptorTypeProgram
DGlcpNAcb1-3DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a3-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(3+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#14: Polysaccharide alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2- ...alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 910.823 Da / Num. of mol.: 2 / Source method: obtained synthetically
DescriptorTypeProgram
DManpa1-3[DManpa1-6]DManpb1-4DGlcpNAcb1-3DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,5,4/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3/a3-b1_b4-c1_c3-d1_c6-e1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(3+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{}[(6+1)][a-D-Manp]{}}}}LINUCSPDB-CARE
#15: Polysaccharide beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-3)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-3DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a3-b1_b4-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(3+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}LINUCSPDB-CARE
#16: Polysaccharide beta-D-mannopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-3DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b3-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(3+1)][b-D-Manp]{}}}LINUCSPDB-CARE
#17: 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.: 22 / Source method: obtained synthetically / Formula: C8H15NO6
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
#18: Sugar...
ChemComp-A2G / 2-acetamido-2-deoxy-alpha-D-galactopyranose / N-acetyl-alpha-D-galactosamine / 2-acetamido-2-deoxy-alpha-D-galactose / 2-acetamido-2-deoxy-D-galactose / 2-acetamido-2-deoxy-galactose / N-ACETYL-2-DEOXY-2-AMINO-GALACTOSE


Type: D-saccharide, alpha linking / Mass: 221.208 Da / Num. of mol.: 24 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGalpNAcaCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-a-D-galactopyranosamineCOMMON NAMEGMML 1.0
a-D-GalpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GalNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 2 types, 90 molecules

#19: Chemical...
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 88 / Source method: obtained synthetically / Formula: Ca
#20: Chemical ChemComp-NI / NICKEL (II) ION


Mass: 58.693 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Ni

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Details

Has ligand of interestN

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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: megalin-RAP complex / Type: COMPLEX / Entity ID: #1-#8 / Source: NATURAL
Source (natural)Organism: Rattus norvegicus (Norway rat)
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen 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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1600 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM software
IDNameVersionCategory
2SerialEMimage acquisition
10RELION3.1.4initial Euler assignment
11RELION3.1.4final Euler assignment
13RELION3.1.43D reconstruction
CTF correctionType: PHASE FLIPPING ONLY
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 67775 / Algorithm: BACK PROJECTION / Symmetry type: POINT

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