[English] 日本語
Yorodumi
- EMDB-15682: Cryo-EM structure of mouse Elp123 with bound SAM -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-15682
TitleCryo-EM structure of mouse Elp123 with bound SAM
Map data
Sample
  • Complex: Mouse Elp123 with bound SAM
    • Protein or peptide: Elongator complex protein 1
    • Protein or peptide: Elongator complex protein 2
    • Protein or peptide: Elongator complex protein 3
  • Ligand: IRON/SULFUR CLUSTERIron–sulfur cluster
  • Ligand: S-ADENOSYLMETHIONINES-Adenosyl methionine
Function / homology
Function and homology information


phosphorylase kinase regulator activity / tRNA uridine(34) acetyltransferase activity / elongator holoenzyme complex / tRNA wobble base 5-methoxycarbonylmethyl-2-thiouridinylation / I-kappaB phosphorylation / tRNA wobble uridine modification / regulation of receptor signaling pathway via JAK-STAT / RNA polymerase II complex binding / acetyltransferase activity / endopeptidase activator activity ...phosphorylase kinase regulator activity / tRNA uridine(34) acetyltransferase activity / elongator holoenzyme complex / tRNA wobble base 5-methoxycarbonylmethyl-2-thiouridinylation / I-kappaB phosphorylation / tRNA wobble uridine modification / regulation of receptor signaling pathway via JAK-STAT / RNA polymerase II complex binding / acetyltransferase activity / endopeptidase activator activity / proteasome assembly / Transferases; Acyltransferases; Transferring groups other than aminoacyl groups / proteasome complex / transcription elongation factor complex / central nervous system development / transcription elongation by RNA polymerase II / neuron migration / : / 4 iron, 4 sulfur cluster binding / tRNA binding / protein kinase activity / positive regulation of cell migration / regulation of transcription by RNA polymerase II / nucleolus / protein kinase binding / nucleoplasm / ATP binding / metal ion binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Elongator complex protein 1 / Elongator complex protein 2 / IKI3 family / Radical SAM, C-terminal extension / Elongator complex protein 3-like / ELP3/YhcC / Radical_SAM C-terminal domain / Elp3/MiaB/NifB / Elongator protein 3, MiaB family, Radical SAM / Radical SAM superfamily ...Elongator complex protein 1 / Elongator complex protein 2 / IKI3 family / Radical SAM, C-terminal extension / Elongator complex protein 3-like / ELP3/YhcC / Radical_SAM C-terminal domain / Elp3/MiaB/NifB / Elongator protein 3, MiaB family, Radical SAM / Radical SAM superfamily / Radical SAM core domain profile. / Radical SAM / Acetyltransferase (GNAT) family / Gcn5-related N-acetyltransferase (GNAT) domain profile. / GNAT domain / Acyl-CoA N-acyltransferase / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Elongator complex protein 1 / Elongator complex protein 2 / Elongator complex protein 3
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.01 Å
AuthorsJaciuk M / Scherf D / Kaszuba K / Gaik M / Koscielniak A / Krutyholowa R / Rawski M / Indyka P / Biela A / Dobosz D ...Jaciuk M / Scherf D / Kaszuba K / Gaik M / Koscielniak A / Krutyholowa R / Rawski M / Indyka P / Biela A / Dobosz D / Lin T-Y / Abbassi N / Hammermeister A / Chramiec-Glabik A / Kosinski J / Schaffrath R / Glatt S
Funding support Poland, European Union, 2 items
OrganizationGrant numberCountry
Polish National Science Centre2018/31/B/NZ1/03559 Poland
European Research Council (ERC)101001394European Union
CitationJournal: Nucleic Acids Res / Year: 2023
Title: Cryo-EM structure of the fully assembled Elongator complex.
Authors: Marcin Jaciuk / David Scherf / Karol Kaszuba / Monika Gaik / Alexander Rau / Anna Kościelniak / Rościsław Krutyhołowa / Michał Rawski / Paulina Indyka / Andrea Graziadei / Andrzej ...Authors: Marcin Jaciuk / David Scherf / Karol Kaszuba / Monika Gaik / Alexander Rau / Anna Kościelniak / Rościsław Krutyhołowa / Michał Rawski / Paulina Indyka / Andrea Graziadei / Andrzej Chramiec-Głąbik / Anna Biela / Dominika Dobosz / Ting-Yu Lin / Nour-El-Hana Abbassi / Alexander Hammermeister / Juri Rappsilber / Jan Kosinski / Raffael Schaffrath / Sebastian Glatt /
Abstract: Transfer RNA (tRNA) molecules are essential to decode messenger RNA codons during protein synthesis. All known tRNAs are heavily modified at multiple positions through post-transcriptional addition ...Transfer RNA (tRNA) molecules are essential to decode messenger RNA codons during protein synthesis. All known tRNAs are heavily modified at multiple positions through post-transcriptional addition of chemical groups. Modifications in the tRNA anticodons are directly influencing ribosome decoding and dynamics during translation elongation and are crucial for maintaining proteome integrity. In eukaryotes, wobble uridines are modified by Elongator, a large and highly conserved macromolecular complex. Elongator consists of two subcomplexes, namely Elp123 containing the enzymatically active Elp3 subunit and the associated Elp456 hetero-hexamer. The structure of the fully assembled complex and the function of the Elp456 subcomplex have remained elusive. Here, we show the cryo-electron microscopy structure of yeast Elongator at an overall resolution of 4.3 Å. We validate the obtained structure by complementary mutational analyses in vitro and in vivo. In addition, we determined various structures of the murine Elongator complex, including the fully assembled mouse Elongator complex at 5.9 Å resolution. Our results confirm the structural conservation of Elongator and its intermediates among eukaryotes. Furthermore, we complement our analyses with the biochemical characterization of the assembled human Elongator. Our results provide the molecular basis for the assembly of Elongator and its tRNA modification activity in eukaryotes.
History
DepositionAug 26, 2022-
Header (metadata) releaseDec 7, 2022-
Map releaseDec 7, 2022-
UpdateMar 29, 2023-
Current statusMar 29, 2023Processing site: PDBe / Status: Released

-
Structure visualization

Supplemental images

emd_15682.png

Downloads & links

-
Map

FileDownload / File: emd_15682.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.86 Å/pix.
x 420 pix.
= 361.2 Å		0.86 Å/pix.
x 420 pix.
= 361.2 Å		0.86 Å/pix.
x 420 pix.
= 361.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.86 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0147
Minimum - Maximum-0.040310964 - 0.1508849
Average (Standard dev.)8.476373e-05 (±0.0012249075)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 361.2 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: #1

Fileemd_15682_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: #2

Fileemd_15682_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Mouse Elp123 with bound SAM

EntireName: Mouse Elp123 with bound SAM
Components
  • Complex: Mouse Elp123 with bound SAM
    • Protein or peptide: Elongator complex protein 1
    • Protein or peptide: Elongator complex protein 2
    • Protein or peptide: Elongator complex protein 3
  • Ligand: IRON/SULFUR CLUSTERIron–sulfur cluster
  • Ligand: S-ADENOSYLMETHIONINES-Adenosyl methionine

-
Supramolecule #1: Mouse Elp123 with bound SAM

SupramoleculeName: Mouse Elp123 with bound SAM / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 610 KDa

-
Macromolecule #1: Elongator complex protein 1

MacromoleculeName: Elongator complex protein 1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 149.756922 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MRNLKLHRTL EFRDIQAPGK PQCFCLRAEQ GTVLIGSERG LTEVDPVRRE VKTEISLVAE GFLPEDGSGC IVGIQDLLDQ ESVCVATAS GDVIVCNLST QQLECVGSVA SGISVMSWSP DQELLLLATA QQTLIMMTKD FEVIAEEQIH QDDFGEGKFV T VGWGSKQT ...String:
MRNLKLHRTL EFRDIQAPGK PQCFCLRAEQ GTVLIGSERG LTEVDPVRRE VKTEISLVAE GFLPEDGSGC IVGIQDLLDQ ESVCVATAS GDVIVCNLST QQLECVGSVA SGISVMSWSP DQELLLLATA QQTLIMMTKD FEVIAEEQIH QDDFGEGKFV T VGWGSKQT QFHGSEGRPT AFPVQLPENA LPWDDRRPHI TWRGDGQYFA VSVVCRQTEA RKIRVWNREF ALQSTSESVP GL GPALAWK PSGSLIASTQ DKPNQQDVVF FEKNGLLHGH FTLPFLKDEV KVNDLLWNAD SSVLAIWLED LPKEDSSTLK SYV QLWTVG NYHWYLKQSL PFSTTGKNQI VSLLWDPVTP CRLHVLCTGW RYLCCDWHWT TDRSSGNSAN DLANVAVIDG NRVL VTVFR QTVVPPPMCT YRLLIPHPVN QVIFSAHLGN DLAVLDASNQ ISVYKCGDKP NMDSTVKLGA VGGNGFKVPL TTPHL EKRY SIQFGNNEEE EEEEVNALQL SFLTWVEDDT FLAISYSHSS SQSIIHHLTV THSEVDEEQG QLDVSSSVTV DGVVIG LCC CSKTKSLAVQ LADGQVLKYL WESPSLAVEP WKNSEGIPVR FVHPCTQMEV ATIGGEECVL GLTDRCRFFI NDTEVAS NI TSFAVCDDFL LVTTHSHTCQ VFSLSGASLK MLQAALSGSH EASGEILRKV ERGSRIVTVV PQDTKLILQM PRGNLEVV H HRALVLAQIR KWLDKLMFKE AFECMRKLRI NLNLIHDHNP KVFLENVETF VKQIDSVNHI NLFFTELREE DVTKTMYPP PITKSVQVST HPDGKKLDLI CDAMRAAMEA INPRKFCLSI LTSHVKKTTP ELEIVLQKVQ ELQGNLPFDP ESVSVEEALK YLLLLVDVN ELFNHSLGTY DFNLVLMVAE KSQKDPKEYL PFLNTLKKME TNYQRFTIDK YLKRYEKALG HLSKCGPEYF T ECLNLIKD KNLYKEALKL YRPDSPQYQA VSMAYGEHLM QEHLYEPAGL VFARCGAQEK ALEAFLACGS WQQALCVAAQ LQ MSKDKVA GLARTLAGKL VEQRKHSEAA TVLEQYAQDY EEAVLLLLEG SAWEEALRLV YKYDRVDIIE TSVKPSILEA QKN YMDFLD SETATFIRHK NRLQVVRALR RQAPQVHVDH EVAHGPESDL FSETSSIMSG SEMSGRYSHS NSRISARSSK NRRK AERKK HSLKEGSPLE GLALLEALSE VVQSVEKLKD EVRAILKVLF LFEFEEQAKE LQRAFESTLQ LMERAVPEIW TPAGQ QSST TPVLGPSSTA NSITASYQQQ KTCVPALDAG VYMPPKMDPR SQWKLSLLE

-
Macromolecule #2: Elongator complex protein 2

MacromoleculeName: Elongator complex protein 2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 93.193773 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MVSSVLEVSH VFCCPNRVRG ALSWNTGPGG LLAFGTSCSV VLYDPQKKVV ITNLNGHTAR VNCLQWIRTE DGSPSNELVS GGSDNRVIH WELENNQVLK SVRLQGHEGP VCAVHAIYQS GPSEGEQHAL IASAASDSTV RIWSKKGSEV KYLQTLSFRD G FVLSVCLA ...String:
MVSSVLEVSH VFCCPNRVRG ALSWNTGPGG LLAFGTSCSV VLYDPQKKVV ITNLNGHTAR VNCLQWIRTE DGSPSNELVS GGSDNRVIH WELENNQVLK SVRLQGHEGP VCAVHAIYQS GPSEGEQHAL IASAASDSTV RIWSKKGSEV KYLQTLSFRD G FVLSVCLA ILPGTNVPVL ACGDDDCRIH LYIQQDDQFQ KALSLCGHED WIRGVEWATF GRDLFLASCS QDCLIRIWRL YM KPASFET KDGSLRLKEN TFTIKDGGVR TTVAVTLETV LAGHENWVNA VHWQPSFYKD GVLQQPVRLL SASMDKTMIL WAP DEESGV WLEQVRVGEV GGNTLGFYDC QFGENGTMII AHAFHGALHL WKQSTVNPRQ WAPEIVISGH FDGVQDLMWD PEGE FIITT STDQTTRLFA PWKKKDQKDR SQVTWHEIAR PQIHGYNIKC LAMIDRFQFV SGADEKVLRV FSAPRNFVEN FSVIS RQSL SHMLCDDQDL PEGATVPALG LSNKALFQGD IASQPFEEDE LISPAFGSPQ VTFQPAVLNE PPTEDHLLQN TLWPEI QKL YGHGYEIVCV ACNNSKTLLA SACKASQKEH AAIILWSTAS WKQVQSLAFH TLTVTQMTFS PDDKFLLAVS RDRTWSL WK RQDATSSEFD PFFSLFAFTN KITSVHSRII WSCDWSPDNK YFFTGSRDKK VVVWGECKSS HNPMEHPIRP CSSILDVG S SVTAVSVCPV LNPAQRYIVA IGLESGKICI YSWNKTNQEI NDWTSCVETN PSQSHSLGIR RLCWKSCSDD DDDDDDDDT EQSEEGPEWL HFASCGEDHT VKIYRVNRRA L

-
Macromolecule #3: Elongator complex protein 3

MacromoleculeName: Elongator complex protein 3 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
EC number: Transferases; Acyltransferases; Transferring groups other than aminoacyl groups
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 62.481059 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MRQKRKGDLS PAELMMLTIG DVIKQLVEAH EQGKDVDLNK MKTKTAAKYG LASQPRLVDI IAAVPPHYRK ILIPKLKAKP VRTASGIAV VAVMCKPHRC PHISFTGNIC IYCPGGPDSD FEYSTQSYTG YEPTSMRAIR ARYDPFLQTR HRIEQLKQLG H SVDKVEFI ...String:
MRQKRKGDLS PAELMMLTIG DVIKQLVEAH EQGKDVDLNK MKTKTAAKYG LASQPRLVDI IAAVPPHYRK ILIPKLKAKP VRTASGIAV VAVMCKPHRC PHISFTGNIC IYCPGGPDSD FEYSTQSYTG YEPTSMRAIR ARYDPFLQTR HRIEQLKQLG H SVDKVEFI VMGGTFMALP EEYRDYFIRS LHDALSGHTS NNIHEAIKYS ERSFTKCVGI TIETRPDYCM KRHLSDMLTY GC TRLEIGV QSVYEDVARD TNRGHTVKAA CESFHLAKDS GFKVVTHMMP DLPNVGLERD IEQFIEFFEN PAFRPDGLKL YPT LVIRGT GLYELWKSGR YRSYSPSDLI ELVARILALV PPWTRVYRVQ RDIPMPLVSS GVEHGNLREL AFARMKDLGI QCRD VRTRE VGIQEIHHRV RPYQVELVRR DYVANGGWET FLSYEDPDQD ILIGLLRLRK CSEETFRFEL GGGVSIVREL HVYGS VVPV SSRDPTKFQH QGFGMLLMEE AERIAREEHG SGKMAVISGV GTRNYYRKIG YRLQGPYMVK MLK

-
Macromolecule #4: IRON/SULFUR CLUSTER

MacromoleculeName: IRON/SULFUR CLUSTER / type: ligand / ID: 4 / Number of copies: 1 / Formula: SF4
Molecular weightTheoretical: 351.64 Da
Chemical component information

ChemComp-FS1:
IRON/SULFUR CLUSTER / Iron–sulfur cluster

-
Macromolecule #5: S-ADENOSYLMETHIONINE

MacromoleculeName: S-ADENOSYLMETHIONINE / type: ligand / ID: 5 / Number of copies: 1 / Formula: SAM
Molecular weightTheoretical: 398.437 Da
Chemical component information

ChemComp-SAM:
S-ADENOSYLMETHIONINE / S-Adenosyl methionine

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration0.4 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
100.0 mMNaClSodium chloridesodium chloride
20.0 mMHEPESHEPES
5.0 mMDithiothreitolDithiothreitol
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: 15 s wait time, blot force 5, 5 s blot time.

-
Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 105000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number real images: 6415 / Average exposure time: 1.82 sec. / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Particle selectionNumber selected: 185923
Details: given number of particles is after TOPAZ picking, 2D cleaning and duplicate removal
Startup modelType of model: NONE
Details: SWISS-MODEL generated full length Elp123 based on 6QK7 structure
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v3.2.0) / Software - details: Ab-Initio Reconstruction / Details: Ab-Initio Reconstruction
Final 3D classificationNumber classes: 3 / Software - Name: RELION (ver. 3.1) / Software - details: masked 3D classification
Details: Masked 3D classification around iron-sulfur cluster
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) / Software - details: 3D auto-refine / Details: 3D auto-refine
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.01 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Software - details: Postprocessing
Details: Declared resolution for the post-process filtered map
Number images used: 42894
Details20 eV slit, fully tuned before the experiment
FSC plot (resolution estimation)

-
Atomic model buiding 1

Initial modelPDB ID:

6qk7

DetailsAfter initial rigid body fit, further fitting was done using NAMDINATOR with manual corrections in COOT
RefinementProtocol: RIGID BODY FIT
Output model

PDB-8avg:
Cryo-EM structure of mouse Elp123 with bound SAM

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more