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- EMDB-33467: Cryo-EM structure of the AKT1-AtKC1 complex from Arabidopsis thaliana -

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Basic information

Entry
Database: EMDB / ID: EMD-33467
TitleCryo-EM structure of the AKT1-AtKC1 complex from Arabidopsis thaliana
Map data
Sample
  • Complex: Arabidopsis thaliana AKT1-AtKC1 complex
    • Protein or peptide: Potassium channel KAT3
    • Protein or peptide: Potassium channel AKT1
  • Ligand: POTASSIUM IONPotassium
Function / homology
Function and homology information


root hair elongation / regulation of stomatal closure / response to nematode / response to water deprivation / inward rectifier potassium channel activity / regulation of monoatomic ion transmembrane transport / monoatomic ion channel complex / potassium ion import across plasma membrane / voltage-gated potassium channel activity / potassium ion transmembrane transport ...root hair elongation / regulation of stomatal closure / response to nematode / response to water deprivation / inward rectifier potassium channel activity / regulation of monoatomic ion transmembrane transport / monoatomic ion channel complex / potassium ion import across plasma membrane / voltage-gated potassium channel activity / potassium ion transmembrane transport / response to salt stress / potassium ion transport / membrane => GO:0016020 / endoplasmic reticulum / identical protein binding / plasma membrane
Similarity search - Function
Potassium channel KAT/AKT / KHA domain / KHA, dimerisation domain of potassium ion channel / KHA domain profile. / Potassium channel, voltage-dependent, EAG/ELK/ERG / Cyclic nucleotide-monophosphate binding domain / Cyclic nucleotide-binding domain / cAMP/cGMP binding motif profile. / Cyclic nucleotide-binding domain / Cyclic nucleotide-binding domain superfamily ...Potassium channel KAT/AKT / KHA domain / KHA, dimerisation domain of potassium ion channel / KHA domain profile. / Potassium channel, voltage-dependent, EAG/ELK/ERG / Cyclic nucleotide-monophosphate binding domain / Cyclic nucleotide-binding domain / cAMP/cGMP binding motif profile. / Cyclic nucleotide-binding domain / Cyclic nucleotide-binding domain superfamily / RmlC-like jelly roll fold / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Potassium channel KAT3 / Potassium channel AKT1
Similarity search - Component
Biological speciesArabidopsis thaliana (thale cress)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsYang GH / Lu YM / Jia YT / Yang F / Zhang YM / Xu X / Li XM / Lei JL
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Commun / Year: 2022
Title: Structural basis for the activity regulation of a potassium channel AKT1 from Arabidopsis.
Authors: Yaming Lu / Miao Yu / Yutian Jia / Fan Yang / Yanming Zhang / Xia Xu / Xiaomin Li / Fan Yang / Jianlin Lei / Yi Wang / Guanghui Yang /
Abstract: The voltage-gated potassium channel AKT1 is responsible for primary K uptake in Arabidopsis roots. AKT1 is functionally activated through phosphorylation and negatively regulated by a potassium ...The voltage-gated potassium channel AKT1 is responsible for primary K uptake in Arabidopsis roots. AKT1 is functionally activated through phosphorylation and negatively regulated by a potassium channel α-subunit AtKC1. However, the molecular basis for the modulation mechanism remains unclear. Here we report the structures of AKT1, phosphorylated-AKT1, a constitutively-active variant, and AKT1-AtKC1 complex. AKT1 is assembled in 2-fold symmetry at the cytoplasmic domain. Such organization appears to sterically hinder the reorientation of C-linkers during ion permeation. Phosphorylated-AKT1 adopts an alternate 4-fold symmetric conformation at cytoplasmic domain, which indicates conformational changes associated with symmetry switch during channel activation. To corroborate this finding, we perform structure-guided mutagenesis to disrupt the dimeric interface and identify a constitutively-active variant Asp379Ala mediates K permeation independently of phosphorylation. This variant predominantly adopts a 4-fold symmetric conformation. Furthermore, the AKT1-AtKC1 complex assembles in 2-fold symmetry. Together, our work reveals structural insight into the regulatory mechanism for AKT1.
History
DepositionMay 18, 2022-
Header (metadata) releaseNov 9, 2022-
Map releaseNov 9, 2022-
UpdateNov 9, 2022-
Current statusNov 9, 2022Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33467.png

Downloads & links

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Map

FileDownload / File: emd_33467.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.86 Å
Density
Contour LevelBy AUTHOR: 0.823
Minimum - Maximum-5.89853 - 8.80325
Average (Standard dev.)0.0119220335 (±0.1830251)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 240.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_33467_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_33467_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Arabidopsis thaliana AKT1-AtKC1 complex

EntireName: Arabidopsis thaliana AKT1-AtKC1 complex
Components
  • Complex: Arabidopsis thaliana AKT1-AtKC1 complex
    • Protein or peptide: Potassium channel KAT3
    • Protein or peptide: Potassium channel AKT1
  • Ligand: POTASSIUM IONPotassium

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Supramolecule #1: Arabidopsis thaliana AKT1-AtKC1 complex

SupramoleculeName: Arabidopsis thaliana AKT1-AtKC1 complex / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293

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Macromolecule #1: Potassium channel KAT3

MacromoleculeName: Potassium channel KAT3 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 75.688023 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MSTTTTEARS PLPLLLRRGR SSTALSASTA EARSPLSILQ FRRRSSKDVR NITSVSSSLL PAFGTFIEDD NPSSKPFIVL HFDRRYRLW ELFLVILVGY SAWASLFELA FEKAAEGALL TIDLVVDFFF AVDIILTFFV SYLDNTTYLN VTDHKLIAKR Y LKSVAFVM ...String:
MSTTTTEARS PLPLLLRRGR SSTALSASTA EARSPLSILQ FRRRSSKDVR NITSVSSSLL PAFGTFIEDD NPSSKPFIVL HFDRRYRLW ELFLVILVGY SAWASLFELA FEKAAEGALL TIDLVVDFFF AVDIILTFFV SYLDNTTYLN VTDHKLIAKR Y LKSVAFVM DVASTLPIQF IYKTITGDVG RGQAFGFLNL LRLWRLRRVA ELFKRLEKDA HFNYFVIRVI KLLCVTIFWI HL AGCILYW IAYHYPRPTD TWIGSQVEDF KERSVWLGYT YSMYWSIVTL TTVGYGDLHA VNSREKTFNM FYMLFNIGLT SYI IGIMTN LVVHGALRTF AMRSAINDIL RYTSKNRLPD TMREQMLAHM QLKFKTAELR QEEVLQDLPK AIRSSINQHL FRSI IEEAY LFKGFPEGLL VQLVSQIQAE YFPPKMEIIL QNEIPTDFYV IVSGGVDIIA SKGVSEQVLA KLGPGSMAGE IGVVF NIPQ PFTVRTRRLS QVIRIGHHKF KEMVQSDNDV DAKMIIANFM TYLKGLNDEL KKEIPFLRDL LDDADAQVQE TVQSEE TPQ SNDEEIVTVS RHENGQIEER RREGVPKRVI IHGQAPPNQD NKNNGDSNGR LIILPDSIQL LFDLAEKKLG KRGSTIA MA DGAHVEQIDA LRENDHLYIF

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Macromolecule #2: Potassium channel AKT1

MacromoleculeName: Potassium channel AKT1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 97.109625 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MRGGALLCGQ VQDEIEQLSR ESSHFSLSTG ILPSLGARSN RRVKLRRFVV SPYDHKYRIW EAFLVVLVVY TAWVSPFEFG FLRKPRPPL SITDNIVNAF FAIDIIMTFF VGYLDKSTYL IVDDRKQIAF KYLRSWFLLD LVSTIPSEAA MRISSQSYGL F NMLRLWRL ...String:
MRGGALLCGQ VQDEIEQLSR ESSHFSLSTG ILPSLGARSN RRVKLRRFVV SPYDHKYRIW EAFLVVLVVY TAWVSPFEFG FLRKPRPPL SITDNIVNAF FAIDIIMTFF VGYLDKSTYL IVDDRKQIAF KYLRSWFLLD LVSTIPSEAA MRISSQSYGL F NMLRLWRL RRVGALFARL EKDRNFNYFW VRCAKLVCVT LFAVHCAACF YYLIAARNSN PAKTWIGANV ANFLEESLWM RY VTSMYWS ITTLTTVGYG DLHPVNTKEM IFDIFYMLFN LGLTAYLIGN MTNLVVHGTS RTRNFRDTIQ AASNFAHRNH LPP RLQDQM LAHLCLKYRT DSEGLQQQET LDALPKAIRS SISHFLFYSL MDKVYLFRGV SNDLLFQLVS EMKAEYFPPK EDVI LQNEA PTDFYILVNG TADLVDVDTG TESIVREVKA GDIIGEIGVL CYRPQLFTVR TKRLCQLLRM NRTTFLNIIQ ANVGD GTII MNNLLQHLKE MNDPVMTNVL LEIENMLARG KMDLPLNLCF AAIREDDLLL HQLLKRGLDP NESDNNGRTP LHIAAS KGT LNCVLLLLEY HADPNCRDAE GSVPLWEAMV EGHEKVVKVL LEHGSTIDAG DVGHFACTAA EQGNLKLLKE IVLHGGD VT RPRATGTSAL HTAVCEENIE MVKYLLEQGA DVNKQDMHGW TPRDLAEQQG HEDIKALFRE KLHERRVHIE TSSSVPIL K TGIRFLGRFT SEPNIRPASR EVSFRIRETR ARRKTNNFDN SLFGILANQS VPKNGLATVD EGRTGNPVRV TISCAEKDD IAGKLVLLPG SFKELLELGS NKFGIVATKV MNKDNNAEID DVDVIRDGDH LIFATDS

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Macromolecule #3: POTASSIUM ION

MacromoleculeName: POTASSIUM ION / type: ligand / ID: 3 / Number of copies: 3 / Formula: K
Molecular weightTheoretical: 39.098 Da

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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

BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1.8 µm / Nominal defocus min: 1.5 µm
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 1.5959 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 104142

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