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- EMDB-22106: High resolution cryoEM structure of huntingtin in complex with HAP40 -

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Basic information

Entry
Database: EMDB / ID: EMD-22106
TitleHigh resolution cryoEM structure of huntingtin in complex with HAP40
Map datarelion local resolution filtered map
Sample
  • Complex: Human huntingtin-HAP40 complex
    • Protein or peptide: Huntingtin
    • Protein or peptide: 40-kDa huntingtin-associated protein
Function / homology
Function and homology information


vesicle cytoskeletal trafficking / regulation of cAMP-dependent protein kinase activity / regulation of phosphoprotein phosphatase activity / positive regulation of inositol 1,4,5-trisphosphate-sensitive calcium-release channel activity / microtubule-based transport / vocal learning / regulation of CAMKK-AMPK signaling cascade / negative regulation of proteasomal protein catabolic process / positive regulation of mitophagy / profilin binding ...vesicle cytoskeletal trafficking / regulation of cAMP-dependent protein kinase activity / regulation of phosphoprotein phosphatase activity / positive regulation of inositol 1,4,5-trisphosphate-sensitive calcium-release channel activity / microtubule-based transport / vocal learning / regulation of CAMKK-AMPK signaling cascade / negative regulation of proteasomal protein catabolic process / positive regulation of mitophagy / profilin binding / vesicle transport along microtubule / positive regulation of cilium assembly / presynaptic cytosol / retrograde vesicle-mediated transport, Golgi to endoplasmic reticulum / positive regulation of aggrephagy / postsynaptic cytosol / positive regulation of lipophagy / dynein intermediate chain binding / beta-tubulin binding / Golgi organization / establishment of mitotic spindle orientation / dynactin binding / Regulation of MECP2 expression and activity / autophagosome / inclusion body / heat shock protein binding / centriole / negative regulation of extrinsic apoptotic signaling pathway / protein destabilization / cytoplasmic vesicle membrane / kinase binding / p53 binding / late endosome / transmembrane transporter binding / early endosome / nuclear body / positive regulation of apoptotic process / axon / apoptotic process / dendrite / perinuclear region of cytoplasm / Golgi apparatus / endoplasmic reticulum / protein-containing complex / nucleoplasm / identical protein binding / nucleus / cytoplasm / cytosol
Similarity search - Function
Factor VIII intron 22 protein / Huntingtin / Huntingtin, middle-repeat / Huntingtin family / : / : / : / Huntingtin, N-terminal HEAT / Huntingtin, bridge / Huntingtin, N-terminal HEAT 1 ...Factor VIII intron 22 protein / Huntingtin / Huntingtin, middle-repeat / Huntingtin family / : / : / : / Huntingtin, N-terminal HEAT / Huntingtin, bridge / Huntingtin, N-terminal HEAT 1 / Huntingtin, C-terminal HEAT / Armadillo-like helical / Tetratricopeptide-like helical domain superfamily / Armadillo-type fold
Similarity search - Domain/homology
40-kDa huntingtin-associated protein / Huntingtin
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsHarding RJ / Deme JC / Lea SM / Arrowsmith CH / Structural Genomics Consortium (SGC)
CitationJournal: Commun Biol / Year: 2021
Title: Huntingtin structure is orchestrated by HAP40 and shows a polyglutamine expansion-specific interaction with exon 1.
Authors: Rachel J Harding / Justin C Deme / Johannes F Hevler / Sem Tamara / Alexander Lemak / Jeffrey P Cantle / Magdalena M Szewczyk / Nola Begeja / Siobhan Goss / Xiaobing Zuo / Peter Loppnau / ...Authors: Rachel J Harding / Justin C Deme / Johannes F Hevler / Sem Tamara / Alexander Lemak / Jeffrey P Cantle / Magdalena M Szewczyk / Nola Begeja / Siobhan Goss / Xiaobing Zuo / Peter Loppnau / Alma Seitova / Ashley Hutchinson / Lixin Fan / Ray Truant / Matthieu Schapira / Jeffrey B Carroll / Albert J R Heck / Susan M Lea / Cheryl H Arrowsmith /
Abstract: Huntington's disease results from expansion of a glutamine-coding CAG tract in the huntingtin (HTT) gene, producing an aberrantly functioning form of HTT. Both wildtype and disease-state HTT form a ...Huntington's disease results from expansion of a glutamine-coding CAG tract in the huntingtin (HTT) gene, producing an aberrantly functioning form of HTT. Both wildtype and disease-state HTT form a hetero-dimer with HAP40 of unknown functional relevance. We demonstrate in vivo and in cell models that HTT and HAP40 cellular abundance are coupled. Integrating data from a 2.6 Å cryo-electron microscopy structure, cross-linking mass spectrometry, small-angle X-ray scattering, and modeling, we provide a near-atomic-level view of HTT, its molecular interaction surfaces and compacted domain architecture, orchestrated by HAP40. Native mass spectrometry reveals a remarkably stable hetero-dimer, potentially explaining the cellular inter-dependence of HTT and HAP40. The exon 1 region of HTT is dynamic but shows greater conformational variety in the polyglutamine expanded mutant than wildtype exon 1. Our data provide a foundation for future functional and drug discovery studies targeting Huntington's disease and illuminate the structural consequences of HTT polyglutamine expansion.
History
DepositionJun 3, 2020-
Header (metadata) releaseJun 17, 2020-
Map releaseJun 17, 2020-
UpdateMar 30, 2022-
Current statusMar 30, 2022Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.035
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.035
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6x9o
  • Surface level: 0.035
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22106.png

Downloads & links

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Map

FileDownload / File: emd_22106.map.gz / Format: CCP4 / Size: 343 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationrelion local resolution filtered map
Voxel sizeX=Y=Z: 0.822 Å
Density
Contour LevelBy AUTHOR: 0.035 / Movie #1: 0.035
Minimum - Maximum-0.11891742 - 0.24614564
Average (Standard dev.)-4.8699287e-05 (±0.0035062376)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions448448448
Spacing448448448
CellA=B=C: 368.256 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8220.8220.822
M x/y/z448448448
origin x/y/z0.0000.0000.000
length x/y/z368.256368.256368.256
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ360360360
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS448448448
D min/max/mean-0.1190.246-0.000

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Supplemental data

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Mask #1

Fileemd_22106_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: relion postprocessed map

Fileemd_22106_additional_1.map
Annotationrelion postprocessed map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: relion auto-refinement map

Fileemd_22106_additional_2.map
Annotationrelion auto-refinement map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: relion auto-refinement unfiltered half map 2

Fileemd_22106_half_map_1.map
Annotationrelion auto-refinement unfiltered half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: relion auto-refinement unfiltered half map 1

Fileemd_22106_half_map_2.map
Annotationrelion auto-refinement unfiltered half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Human huntingtin-HAP40 complex

EntireName: Human huntingtin-HAP40 complex
Components
  • Complex: Human huntingtin-HAP40 complex
    • Protein or peptide: Huntingtin
    • Protein or peptide: 40-kDa huntingtin-associated protein

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Supramolecule #1: Human huntingtin-HAP40 complex

SupramoleculeName: Human huntingtin-HAP40 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)

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Macromolecule #1: Huntingtin

MacromoleculeName: Huntingtin / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 349.472344 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MATLEKLMKA FESLKSFQQQ QQQQQQQQQQ QQQQQQQQQQ PPPPPPPPPP PQLPQPPPQA QPLLPQPQPP PPPPPPPPGP AVAEEPLHR PKKELSATKK DRVNHCLTIC ENIVAQSVRN SPEFQKLLGI AMELFLLCSD DAESDVRMVA DECLNKVIKA L MDSNLPRL ...String:
MATLEKLMKA FESLKSFQQQ QQQQQQQQQQ QQQQQQQQQQ PPPPPPPPPP PQLPQPPPQA QPLLPQPQPP PPPPPPPPGP AVAEEPLHR PKKELSATKK DRVNHCLTIC ENIVAQSVRN SPEFQKLLGI AMELFLLCSD DAESDVRMVA DECLNKVIKA L MDSNLPRL QLELYKEIKK NGAPRSLRAA LWRFAELAHL VRPQKCRPYL VNLLPCLTRT SKRPEESVQE TLAAAVPKIM AS FGNFAND NEIKVLLKAF IANLKSSSPT IRRTAAGSAV SICQHSRRTQ YFYSWLLNVL LGLLVPVEDE HSTLLILGVL LTL RYLVPL LQQQVKDTSL KGSFGVTRKE MEVSPSAEQL VQVYELTLHH TQHQDHNVVT GALELLQQLF RTPPPELLQT LTAV GGIGQ LTAAKEESGG RSRSGSIVEL IAGGGSSCSP VLSRKQKGKV LLGEEEALED DSESRSDVSS SALTASVKDE ISGEL AASS GVSTPGSAGH DIITEQPRSQ HTLQADSVDL ASCDLTSSAT DGDEEDILSH SSSQVSAVPS DPAMDLNDGT QASSPI SDS SQTTTEGPDS AVTPSDSSEI VLDGTDNQYL GLQIGQPQDE DEEATGILPD EASEAFRNSS MALQQAHLLK NMSHCRQ PS DSSVDKFVLR DEATEPGDQE NKPCRIKGDI GQSTDDDSAP LVHCVRLLSA SFLLTGGKNV LVPDRDVRVS VKALALSC V GAAVALHPES FFSKLYKVPL DTTEYPEEQY VSDILNYIDH GDPQVRGATA ILCGTLICSI LSRSRFHVGD WMGTIRTLT GNTFSLADCI PLLRKTLKDE SSVTCKLACT AVRNCVMSLC SSSYSELGLQ LIIDVLTLRN SSYWLVRTEL LETLAEIDFR LVSFLEAKA ENLHRGAHHY TGLLKLQERV LNNVVIHLLG DEDPRVRHVA AASLIRLVPK LFYKCDQGQA DPVVAVARDQ S SVYLKLLM HETQPPSHFS VSTITRIYRG YNLLPSITDV TMENNLSRVI AAVSHELITS TTRALTFGCC EALCLLSTAF PV CIWSLGW HCGVPPLSAS DESRKSCTVG MATMILTLLS SAWFPLDLSA HQDALILAGN LLAASAPKSL RSSWASEEEA NPA ATKQEE VWPALGDRAL VPMVEQLFSH LLKVINICAH VLDDVAPGPA IKAALPSLTN PPSLSPIRRK GKEKEPGEQA SVPL SPKKG SEASAASRQS DTSGPVTTSK SSSLGSFYHL PSYLKLHDVL KATHANYKVT LDLQNSTEKF GGFLRSALDV LSQIL ELAT LQDIGKCVEE ILGYLKSCFS REPMMATVCV QQLLKTLFGT NLASQFDGLS SNPSKSQGRA QRLGSSSVRP GLYHYC FMA PYTHFTQALA DASLRNMVQA EQENDTSGWF DVLQKVSTQL KTNLTSVTKN RADKNAIHNH IRLFEPLVIK ALKQYTT TT CVQLQKQVLD LLAQLVQLRV NYCLLDSDQV FIGFVLKQFE YIEVGQFRES EAIIPNIFFF LVLLSYERYH SKQIIGIP K IIQLCDGIMA SGRKAVTHAI PALQPIVHDL FVLRGTNKAD AGKELETQKE VVVSMLLRLI QYHQVLEMFI LVLQQCHKE NEDKWKRLSR QIADIILPML AKQQMHIDSH EALGVLNTLF EILAPSSLRP VDMLLRSMFV TPNTMASVST VQLWISGILA ILRVLISQS TEDIVLSRIQ ELSFSPYLIS CTVINRLRDG DSTSTLEEHS EGKQIKNLPE ETFSRFLLQL VGILLEDIVT K QLKVEMSE QQHTFYCQEL GTLLMCLIHI FKSGMFRRIT AAATRLFRSD GCGGSFYTLD SLNLRARSMI TTHPALVLLW CQ ILLLVNH TDYRWWAEVQ QTPKRHSLSS TKLLSPQMSG EEEDSDLAAK LGMCNREIVR RGALILFCDY VCQNLHDSEH LTW LIVNHI QDLISLSHEP PVQDFISAVH RNSAASGLFI QAIQSRCENL STPTMLKKTL QCLEGIHLSQ SGAVLTLYVD RLLC TPFRV LARMVDILAC RRVEMLLAAN LQSSMAQLPM EELNRIQEYL QSSGLAQRHQ RLYSLLDRFR LSTMQDSLSP SPPVS SHPL DGDGHVSLET VSPDKDWYVH LVKSQCWTRS DSALLEGAEL VNRIPAEDMN AFMMNSEFNL SLLAPCLSLG MSEISG GQK SALFEAAREV TLARVSGTVQ QLPAVHHVFQ PELPAEPAAY WSKLNDLFGD AALYQSLPTL ARALAQYLVV VSKLPSH LH LPPEKEKDIV KFVVATLEAL SWHLIHEQIP LSLDLQAGLD CCCLALQLPG LWSVVSSTEF VTHACSLIHC VHFILEAV A VQPGEQLLSP ERRTNTPKAI SEEEEEVDPN TQNPKYITAA CEMVAEMVES LQSVLALGHK RNSGVPAFLT PLLRNIIIS LARLPLVNSY TRVPPLVWKL GWSPKPGGDF GTAFPEIPVE FLQEKEVFKE FIYRINTLGW TSRTQFEETW ATLLGVLVTQ PLVMEQEES PPEEDTERTQ INVLAVQAIT SLVLSAMTVP VAGNPAVSCL EQQPRNKPLK ALDTRFGRKL SIIRGIVEQE I QAMVSKRE NIATHHLYQA WDPVPSLSPA TTGALISHEK LLLQINPERE LGSMSYKLGQ VSIHSVWLGN SITPLREEEW DE EEEEEAD APAPSSPPTS PVNSRKHRAG VDIHSCSQFL LELYSRWILP SSSARRTPAI LISEVVRSLL VVSDLFTERN QFE LMYVTL TELRRVHPSE DEILAQYLVP ATCKAAAVLG MDKAVAEPVS RLLESTLRSS HLPSRVGALH GILYVLECDL LDDT AKQLI PVISDYLLSN LKGIAHCVNI HSQQHVLVMC ATAFYLIENY PLDVGPEFSA SIIQMCGVML SGSEESTPSI IYHCA LRGL ERLLLSEQLS RLDAESLVKL SVDRVNVHSP HRAMAALGLM LTCMYTGKEK VSPGRTSDPN PAAPDSESVI VAMERV SVL FDRIRKGFPC EARVVARILP QFLDDFFPPQ DIMNKVIGEF LSNQQPYPQF MATVVYKVFQ TLHSTGQSSM VRDWVML SL SNFTQRAPVA MATWSLSCFF VSASTSPWVA AILPHVISRM GKLEQVDVNL FCLVATDFYR HQIEEELDRR AFQSVLEV V AAPGSPYHRL LTCLRNVHKV TTCGGSGDYK DDDDK

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Macromolecule #2: 40-kDa huntingtin-associated protein

MacromoleculeName: 40-kDa huntingtin-associated protein / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 41.342254 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MHHHHHHSSG RENLYFQGMA AAAAGLGGGG AGPGPEAGDF LARYRLVSNK LKKRFLRKPN VAEAGEQFGQ LGRELRAQEC LPYAAWCQL AVARCQQALF HGPGEALALT EAARLFLRQE RDARQRLVCP AAYGEPLQAA ASALGAAVRL HLELGQPAAA A ALCLELAA ...String:
MHHHHHHSSG RENLYFQGMA AAAAGLGGGG AGPGPEAGDF LARYRLVSNK LKKRFLRKPN VAEAGEQFGQ LGRELRAQEC LPYAAWCQL AVARCQQALF HGPGEALALT EAARLFLRQE RDARQRLVCP AAYGEPLQAA ASALGAAVRL HLELGQPAAA A ALCLELAA ALRDLGQPAA AAGHFQRAAQ LQLPQLPLAA LQALGEAASC QLLARDYTGA LAVFTRMQRL AREHGSHPVQ SL PPPPPPA PQPGPGATPA LPAALLPPNS GSAAPSPAAL GAFSDVLVRC EVSRVLLLLL LQPPPAKLLP EHAQTLEKYS WEA FDSHGQ ESSGQLPEEL FLLLQSLVMA THEKDTEAIK SLQVEMWPLL TAEQNHLLHL VLQETISPSG QGV

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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

Concentration0.2 mg/mL
BufferpH: 7.4
Component:
ConcentrationNameFormula
25.0 mMHEPES
300.0 mMNaCl
1.0 mMDTT
0.025 % v/vCHAPS
GridModel: Quantifoil R2/4 / Material: GOLD / Mesh: 300 / Support film - Material: GRAPHENE / Support film - topology: CONTINUOUS / Support film - Film thickness: 0.35000000000000003 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Details: 15 mA
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Average exposure time: 8.0 sec. / Average electron dose: 48.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 165000
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

CTF correctionSoftware - Name: SIMPLE (ver. 3.0b)
Startup modelType of model: OTHER / Details: relion SGD de novo map from particles
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 647468
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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