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- EMDB-45209: AP-3 Arf1 dimeric interface, focused refinement -

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Basic information

Entry
Database: EMDB / ID: EMD-45209
TitleAP-3 Arf1 dimeric interface, focused refinement
Map datadeepEMHancer sharpened map for Arf1 dimer
Sample
  • Complex: AP-3 Arf1 dimeric interface, focused refinement
    • Protein or peptide: AP-3 complex subunit beta-1
    • Protein or peptide: AP-3 complex subunit mu-1
    • Protein or peptide: ADP-ribosylation factor 1
    • Protein or peptide: Lysosome-associated membrane glycoprotein 1
  • Ligand: MAGNESIUM ION
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
KeywordsAdaptor Protein complex / Endosomal Trafficking / Lysosomal Trafficking / Protein transport / AP complex / TRANSPORT PROTEIN
Function / homology
Function and homology information


establishment of protein localization to mitochondrial membrane involved in mitochondrial fission / clathrin-coated vesicle cargo loading, AP-3-mediated / skin epidermis development / regulation of organelle transport along microtubule / AP-3 adaptor complex / positive regulation of natural killer cell degranulation / granzyme-mediated programmed cell death signaling pathway / anterograde synaptic vesicle transport / microvesicle / phagolysosome membrane ...establishment of protein localization to mitochondrial membrane involved in mitochondrial fission / clathrin-coated vesicle cargo loading, AP-3-mediated / skin epidermis development / regulation of organelle transport along microtubule / AP-3 adaptor complex / positive regulation of natural killer cell degranulation / granzyme-mediated programmed cell death signaling pathway / anterograde synaptic vesicle transport / microvesicle / phagolysosome membrane / Golgi to lysosome transport / mitotic cleavage furrow ingression / cytolytic granule membrane / trans-Golgi Network Vesicle Budding / establishment of protein localization to organelle / postsynaptic recycling endosome / clathrin adaptor complex / platelet dense granule organization / Glycosphingolipid transport / regulation of receptor internalization / melanosome assembly / granulocyte differentiation / Intra-Golgi traffic / regulation of Arp2/3 complex-mediated actin nucleation / postsynaptic neurotransmitter receptor internalization / positive regulation of NK T cell differentiation / GTP-dependent protein binding / Synthesis of PIPs at the Golgi membrane / lysosomal lumen acidification / clathrin-coated vesicle membrane / positive regulation of natural killer cell mediated cytotoxicity / antigen processing and presentation, exogenous lipid antigen via MHC class Ib / protein targeting to lysosome / respiratory system process / melanosome organization / anterograde axonal transport / intracellular zinc ion homeostasis / Nef Mediated CD4 Down-regulation / dendritic spine organization / protein localization to cell surface / long-term synaptic depression / COPI-dependent Golgi-to-ER retrograde traffic / azurophil granule membrane / Lysosome Vesicle Biogenesis / toll-like receptor signaling pathway / ion channel inhibitor activity / Golgi Associated Vesicle Biogenesis / lysosome organization / lung morphogenesis / cell leading edge / Association of TriC/CCT with target proteins during biosynthesis / autolysosome / Synthesis of PIPs at the plasma membrane / ficolin-1-rich granule membrane / autophagosome membrane / homeostasis of number of cells / intracellular copper ion homeostasis / intracellular transport / single fertilization / hematopoietic progenitor cell differentiation / COPI-mediated anterograde transport / vesicle-mediated transport / axon cytoplasm / MHC class II antigen presentation / multivesicular body / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / sarcomere / small monomeric GTPase / intracellular protein transport / mRNA transcription by RNA polymerase II / cytoplasmic vesicle membrane / protein modification process / sarcolemma / cell morphogenesis / cellular response to virus / small GTPase binding / endocytosis / blood coagulation / Signaling by BRAF and RAF1 fusions / late endosome membrane / melanosome / late endosome / synaptic vesicle / virus receptor activity / protein phosphatase binding / cytoplasmic vesicle / spermatogenesis / early endosome / lysosome / protein stabilization / endosome membrane / postsynaptic density / neuron projection / inflammatory response / protein domain specific binding / lysosomal membrane / Golgi membrane / external side of plasma membrane / focal adhesion / GTPase activity
Similarity search - Function
AP-3 complex subunit beta, C-terminal domain / AP-3 complex subunit beta 1, serine-rich domain / Clathrin-adaptor complex-3 beta-1 subunit C-terminal / Serine-rich region of AP3B1, clathrin-adaptor complex / Clathrin-adaptor complex-3 beta-1 subunit C-terminal / Lysosome-associated membrane glycoprotein, conserved site / AP-3 complex subunit beta / : / Lysosome-associated membrane glycoprotein 2, transmembrane segment / Lysosome-associated membrane glycoproteins duplicated domain signature. ...AP-3 complex subunit beta, C-terminal domain / AP-3 complex subunit beta 1, serine-rich domain / Clathrin-adaptor complex-3 beta-1 subunit C-terminal / Serine-rich region of AP3B1, clathrin-adaptor complex / Clathrin-adaptor complex-3 beta-1 subunit C-terminal / Lysosome-associated membrane glycoprotein, conserved site / AP-3 complex subunit beta / : / Lysosome-associated membrane glycoprotein 2, transmembrane segment / Lysosome-associated membrane glycoproteins duplicated domain signature. / LAMP glycoproteins transmembrane and cytoplasmic domain signature. / Lysosome-associated membrane glycoprotein / : / Lysosome-associated membrane glycoprotein 2-like, luminal domains / Lysosome-associated membrane glycoprotein family profile. / ADP-ribosylation factor 1-5 / Beta-adaptin appendage, C-terminal subdomain / Beta2-adaptin appendage, C-terminal sub-domain / AP complex subunit beta / Clathrin adaptor complexes medium chain signature 1. / Clathrin adaptor, mu subunit / Clathrin adaptor, mu subunit, conserved site / Clathrin adaptor complexes medium chain signature 2. / : / Adaptor complexes medium subunit family / AP complex, mu/sigma subunit / Clathrin adaptor complex small chain / AP-2 complex subunit mu, C-terminal superfamily / Mu homology domain / Mu homology domain (MHD) profile. / Clathrin/coatomer adaptor, adaptin-like, N-terminal / Adaptin N terminal region / Small GTPase superfamily, ARF type / small GTPase Arf family profile. / Sar1p-like members of the Ras-family of small GTPases / Small GTPase superfamily, ARF/SAR type / ADP-ribosylation factor family / Longin-like domain superfamily / ARF-like small GTPases; ARF, ADP-ribosylation factor / Rab subfamily of small GTPases / Armadillo-like helical / Small GTP-binding protein domain / Armadillo-type fold / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
AP-3 complex subunit beta-1 / Lysosome-associated membrane glycoprotein 1 / ADP-ribosylation factor 1 / AP-3 complex subunit mu-1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.2 Å
AuthorsBegley MC / Baker RW
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM150960 United States
CitationJournal: To Be Published
Title: A structure-based mechanism for initiation of AP-3 coated vesicle formation
Authors: Begley M / Baker RW
History
DepositionJun 6, 2024-
Header (metadata) releaseDec 18, 2024-
Map releaseDec 18, 2024-
UpdateDec 18, 2024-
Current statusDec 18, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45209.png

Downloads & links

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Map

FileDownload / File: emd_45209.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationdeepEMHancer sharpened map for Arf1 dimer
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 400 pix.
= 423.2 Å		1.06 Å/pix.
x 400 pix.
= 423.2 Å		1.06 Å/pix.
x 400 pix.
= 423.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.058 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.0017204892 - 2.0888836
Average (Standard dev.)0.0008835663 (±0.021501599)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 423.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_45209_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Mask #2

Fileemd_45209_msk_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Additional map: Unsharpened map for Arf1 dimer

Fileemd_45209_additional_1.map
AnnotationUnsharpened map for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Sharpened map for Arf1 dimer

Fileemd_45209_additional_2.map
AnnotationSharpened map for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Mask for Arf1 dimer

Fileemd_45209_additional_3.map
AnnotationMask for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Refined mask for Arf1 dimer

Fileemd_45209_additional_4.map
AnnotationRefined mask for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Sharpened map for Arf1 dimer

Fileemd_45209_additional_5.map
AnnotationSharpened map for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Map for filtering by local resoluion for Arf1 dimer

Fileemd_45209_additional_6.map
AnnotationMap for filtering by local resoluion for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half map for Arf1 dimer

Fileemd_45209_half_map_1.map
AnnotationUnfiltered half map for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half map for Arf1 dimer

Fileemd_45209_half_map_2.map
AnnotationUnfiltered half map for Arf1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : AP-3 Arf1 dimeric interface, focused refinement

EntireName: AP-3 Arf1 dimeric interface, focused refinement
Components
  • Complex: AP-3 Arf1 dimeric interface, focused refinement
    • Protein or peptide: AP-3 complex subunit beta-1
    • Protein or peptide: AP-3 complex subunit mu-1
    • Protein or peptide: ADP-ribosylation factor 1
    • Protein or peptide: Lysosome-associated membrane glycoprotein 1
  • Ligand: MAGNESIUM ION
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE

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Supramolecule #1: AP-3 Arf1 dimeric interface, focused refinement

SupramoleculeName: AP-3 Arf1 dimeric interface, focused refinement / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Molecular weightTheoretical: 256 KDa

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Macromolecule #1: AP-3 complex subunit beta-1

MacromoleculeName: AP-3 complex subunit beta-1 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 77.286523 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSSNSFPYNE QSGGGEATEL GQEATSTISP SGAFGLFSSD LKKNEDLKQM LESNKDSAKL DAMKRIVGMI AKGKNASELF PAVVKNVAS KNIEIKKLVY VYLVRYAEEQ QDLALLSIST FQRALKDPNQ LIRASALRVL SSIRVPIIVP IMMLAIKEAS A DLSPYVRK ...String:
MSSNSFPYNE QSGGGEATEL GQEATSTISP SGAFGLFSSD LKKNEDLKQM LESNKDSAKL DAMKRIVGMI AKGKNASELF PAVVKNVAS KNIEIKKLVY VYLVRYAEEQ QDLALLSIST FQRALKDPNQ LIRASALRVL SSIRVPIIVP IMMLAIKEAS A DLSPYVRK NAAHAIQKLY SLDPEQKEML IEVIEKLLKD KSTLVAGSVV MAFEEVCPDR IDLIHKNYRK LCNLLVDVEE WG QVVIIHM LTRYARTQFV SPWKEGDELE DNGKNFYESD DDQKEKTDKK KKPYTMDPDH RLLIRNTKPL LQSRNAAVVM AVA QLYWHI SPKSEAGIIS KSLVRLLRSN REVQYIVLQN IATMSIQRKG MFEPYLKSFY VRSTDPTMIK TLKLEILTNL ANEA NISTL LREFQTYVKS QDKQFAAATI QTIGRCATNI LEVTDTCLNG LVCLLSNRDE IVVAESVVVI KKLLQMQPAQ HGEII KHMA KLLDSITVPV ARASILWLIG ENCERVPKIA PDVLRKMAKS FTSEDDLVKL QILNLGAKLY LTNSKQTKLL TQYILN LGK YDQNYDIRDR TRFIRQLIVP NVKSGALSKY AKKIFLAQKP APLLESPFKD RDHFQLGTLS HTLNIKATGY LELSNWP EV APDPSVRNVE VIELAKEWTP AGKAKQENSA KKFYSGLEVL FQ

UniProtKB: AP-3 complex subunit beta-1

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Macromolecule #2: AP-3 complex subunit mu-1

MacromoleculeName: AP-3 complex subunit mu-1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 46.989965 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MIHSLFLINC SGDIFLEKHW KSVVSQSVCD YFFEAQEKAA DVENVPPVIS TPHHYLISIY RDKLFFVSVI QTEVPPLFVI EFLHRVADT FQDYFGECSE AAIKDNVVIV YELLEEMLDN GFPLATESNI LKELIKPPTI LRSVVNSITG SSNVGDTLPT G QLSNIPWR ...String:
MIHSLFLINC SGDIFLEKHW KSVVSQSVCD YFFEAQEKAA DVENVPPVIS TPHHYLISIY RDKLFFVSVI QTEVPPLFVI EFLHRVADT FQDYFGECSE AAIKDNVVIV YELLEEMLDN GFPLATESNI LKELIKPPTI LRSVVNSITG SSNVGDTLPT G QLSNIPWR RAGVKYTNNE AYFDVVEEID AIIDKSGSTV FAEIQGVIDA CIKLSGMPDL SLSFMNPRLL DDVSFHPCIR FK RWESERV LSFIPPDGNF RLISYRVSSQ NLVAIPVYVK HSISFKENSS CGRFDITIGP KQNMGKTIEG ITVTVHMPKV VLN MNLTPT QGSYTFDPVT KVLTWDVGKI TPQKLPSLKG LVNLQSGAPK PEENPSLNIQ FKIQQLAISG LKVNRLDMYG EKYK PFKGV KYVTKAGKFQ VRT

UniProtKB: AP-3 complex subunit mu-1

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Macromolecule #3: ADP-ribosylation factor 1

MacromoleculeName: ADP-ribosylation factor 1 / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO / EC number: small monomeric GTPase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 20.775812 KDa
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString:
GNIFANLFKG LFGKKEMRIL MVGLDAAGKT TILYKLKLGE IVTTIPTIGF NVETVEYKNI SFTVWDVGGL DKIRPLWRHY FQNTQGLIF VVDSNDRERV NEAREELMRM LAEDELRDAV LLVFANKQDL PNAMNAAEIT DKLGLHSLRH RNWYIQATCA T SGDGLYEG LDWLSNQLRN QKSL

UniProtKB: ADP-ribosylation factor 1

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Macromolecule #4: Lysosome-associated membrane glycoprotein 1

MacromoleculeName: Lysosome-associated membrane glycoprotein 1 / type: protein_or_peptide / ID: 4 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 1.377553 KDa
SequenceString:
GRKRSHAGYQ TI

UniProtKB: Lysosome-associated membrane glycoprotein 1

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Macromolecule #5: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #6: GUANOSINE-5'-TRIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 6 / Number of copies: 2 / Formula: GTP
Molecular weightTheoretical: 523.18 Da
Chemical component information

ChemComp-GTP:
GUANOSINE-5'-TRIPHOSPHATE / GTP, energy-carrying molecule*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
2.7 mMKClPotassium Chloride
10.0 mMNa2HPO4Sodium Phosphate (Dibasic)
1.8 mMKH2PO4Potassium Phosphate (Monobasic)
150.0 mMNaClSodium Chloride
1.0 mMC9H15O6PTCEP
5.0 mMC10H16N2O8EDTA
10.0 mMMgCl2Magnesium Chloride
2.0 mMC10H16N5O14P3GTP

Details: 1x PBS (pH 7.4), 150mM NaCl, 1mM TCEP, 2mM GTP, 5mM EDTA, 10mM MgCl2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 200 sec. / Pretreatment - Atmosphere: AIR
Details: Quantifoil Active grids (SPT Labtech) with backside gold coated. Plasma was 12mA for cleaning
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 294 K / Instrument: SPOTITON / Details: Commercial form of the chameleon (SPT Labtech).
DetailsAP-3 dimer bound to myristoylated Arf1 (Q71L) and LAMP1 on a lipid nanodisc; focused refinement on Arf1 dimeric interface

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 53.37 e/Å2
Details: 2 datasets collected independently and merged for processing
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.4000000000000001 µm / Nominal defocus min: 0.4 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.4.1) / Number images used: 122155
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.4.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.4.1)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 4.4.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementProtocol: AB INITIO MODEL
Output model

PDB-9c5a:
AP-3 Arf1 dimeric interface, focused refinement

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