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- EMDB-28864: Human ABCA4 structure in complex with AMP-PNP -

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Basic information

Entry
Database: EMDB / ID: EMD-28864
TitleHuman ABCA4 structure in complex with AMP-PNP
Map data
Sample
  • Complex: ABCA4 structure in complex with AMP-PNP
    • Protein or peptide: Retinal-specific phospholipid-transporting ATPase ABCA4
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: MAGNESIUM ION
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
KeywordsABC transporter / importer / membrane protein / TRANSPORT PROTEIN
Function / homology
Function and homology information


rod photoreceptor disc membrane / flippase activity / N-retinylidene-phosphatidylethanolamine flippase activity / all-trans retinal binding / phospholipid transfer to membrane / retinol transmembrane transporter activity / phospholipid transporter activity / 11-cis retinal binding / retinal metabolic process / ATPase-coupled intramembrane lipid transporter activity ...rod photoreceptor disc membrane / flippase activity / N-retinylidene-phosphatidylethanolamine flippase activity / all-trans retinal binding / phospholipid transfer to membrane / retinol transmembrane transporter activity / phospholipid transporter activity / 11-cis retinal binding / retinal metabolic process / ATPase-coupled intramembrane lipid transporter activity / phosphatidylethanolamine flippase activity / phototransduction, visible light / retinoid binding / P-type phospholipid transporter / photoreceptor cell maintenance / phospholipid translocation / retinoid metabolic process / lipid transport / Retinoid cycle disease events / The canonical retinoid cycle in rods (twilight vision) / ATPase-coupled transmembrane transporter activity / photoreceptor outer segment / ABC-type transporter activity / visual perception / ABC-family proteins mediated transport / transmembrane transport / photoreceptor disc membrane / cytoplasmic vesicle / intracellular membrane-bounded organelle / GTPase activity / endoplasmic reticulum / ATP hydrolysis activity / ATP binding / membrane / plasma membrane
Similarity search - Function
Retinal-specific ATP-binding cassette transporter / ABC transporter A / ABC-2 family transporter protein / ABC-2 type transporter / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / ATPases associated with a variety of cellular activities ...Retinal-specific ATP-binding cassette transporter / ABC transporter A / ABC-2 family transporter protein / ABC-2 type transporter / ABC transporter-like, conserved site / ABC transporters family signature. / ABC transporter / ABC transporter-like, ATP-binding domain / ATP-binding cassette, ABC transporter-type domain profile. / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Retinal-specific phospholipid-transporting ATPase ABCA4
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.95 Å
AuthorsScortecci JF / Van Petegem F / Molday RS
Funding support Canada, 1 items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR)PJT175118 Canada
CitationJournal: J Biol Chem / Year: 2024
Title: Structural and functional characterization of the nucleotide-binding domains of ABCA4 and their role in Stargardt disease.
Authors: Jessica Fernandes Scortecci / Fabian A Garces / Jai K Mahto / Laurie L Molday / Filip Van Petegem / Robert S Molday /
Abstract: ABCA4 is an ATP-binding cassette (ABC) transporter that prevents the buildup of toxic retinoid compounds by facilitating the transport of N-retinylidene-phosphatidylethanolamine across membranes of ...ABCA4 is an ATP-binding cassette (ABC) transporter that prevents the buildup of toxic retinoid compounds by facilitating the transport of N-retinylidene-phosphatidylethanolamine across membranes of rod and cone photoreceptor cells. Over 1500 missense mutations in ABCA4, many in the nucleotide-binding domains (NBDs), have been genetically linked to Stargardt disease. Here, we show by cryo-EM that ABCA4 is converted from an open outward conformation to a closed conformation upon the binding of adenylyl-imidodiphosphate. Structural information and biochemical studies were used to further define the role of the NBDs in the functional properties of ABCA4 and the mechanisms by which mutations lead to the loss in activity. We show that ATPase activity in both NBDs is required for the functional activity of ABCA4. Mutations in Walker A asparagine residues cause a severe reduction in substrate-activated ATPase activity due to the loss in polar interactions with residues within the D-loops of the opposing NBD. The structural basis for how disease mutations in other NBD residues, including the R1108C, R2077W, R2107H, and L2027F, affect the structure and function of ABCA4 is described. Collectively, our studies provide insight into the structure and function of ABCA4 and mechanisms underlying Stargardt disease.
History
DepositionNov 13, 2022-
Header (metadata) releaseNov 22, 2023-
Map releaseNov 22, 2023-
UpdateSep 18, 2024-
Current statusSep 18, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_28864.png

Downloads & links

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Map

FileDownload / File: emd_28864.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 360 pix.
= 377.65 Å		1.05 Å/pix.
x 360 pix.
= 377.65 Å		1.05 Å/pix.
x 360 pix.
= 377.65 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.04903 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.308
Minimum - Maximum-0.48872125 - 1.1000237
Average (Standard dev.)0.0005532081 (±0.031552333)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 377.65 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_28864_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_28864_half_map_2.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Sample components

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Entire : ABCA4 structure in complex with AMP-PNP

EntireName: ABCA4 structure in complex with AMP-PNP
Components
  • Complex: ABCA4 structure in complex with AMP-PNP
    • Protein or peptide: Retinal-specific phospholipid-transporting ATPase ABCA4
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: MAGNESIUM ION
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER

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Supramolecule #1: ABCA4 structure in complex with AMP-PNP

SupramoleculeName: ABCA4 structure in complex with AMP-PNP / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: Recombinantly expressed human ABCA4
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 250 KDa

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Macromolecule #1: Retinal-specific phospholipid-transporting ATPase ABCA4

MacromoleculeName: Retinal-specific phospholipid-transporting ATPase ABCA4
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: P-type phospholipid transporter
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 256.202531 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGFVRQIQLL LWKNWTLRKR QKIRFVVELV WPLSLFLVLI WLRNANPLYS HHECHFPNKA MPSAGMLPWL QGIFCNVNNP CFQSPTPGE SPGIVSNYNN SILARVYRDF QELLMNAPES QHLGRIWTEL HILSQFMDTL RTHPERIAGR GIRIRDILKD E ETLTLFLI ...String:
MGFVRQIQLL LWKNWTLRKR QKIRFVVELV WPLSLFLVLI WLRNANPLYS HHECHFPNKA MPSAGMLPWL QGIFCNVNNP CFQSPTPGE SPGIVSNYNN SILARVYRDF QELLMNAPES QHLGRIWTEL HILSQFMDTL RTHPERIAGR GIRIRDILKD E ETLTLFLI KNIGLSDSVV YLLINSQVRP EQFAHGVPDL ALKDIACSEA LLERFIIFSQ RRGAKTVRYA LCSLSQGTLQ WI EDTLYAN VDFFKLFRVL PTLLDSRSQG INLRSWGGIL SDMSPRIQEF IHRPSMQDLL WVTRPLMQNG GPETFTKLMG ILS DLLCGY PEGGGSRVLS FNWYEDNNYK AFLGIDSTRK DPIYSYDRRT TSFCNALIQS LESNPLTKIA WRAAKPLLMG KILY TPDSP AARRILKNAN STFEELEHVR KLVKAWEEVG PQIWYFFDNS TQMNMIRDTL GNPTVKDFLN RQLGEEGITA EAILN FLYK GPRESQADDM ANFDWRDIFN ITDRTLRLVN QYLECLVLDK FESYNDETQL TQRALSLLEE NMFWAGVVFP DMYPWT SSL PPHVKYKIRM DIDVVEKTNK IKDRYWDSGP RADPVEDFRY IWGGFAYLQD MVEQGITRSQ VQAEAPVGIY LQQMPYP CF VDDSFMIILN RCFPIFMVLA WIYSVSMTVK SIVLEKELRL KETLKNQGVS NAVIWCTWFL DSFSIMSMSI FLLTIFIM H GRILHYSDPF ILFLFLLAFS TATIMLCFLL STFFSKASLA AACSGVIYFT LYLPHILCFA WQDRMTAELK KAVSLLSPV AFGFGTEYLV RFEEQGLGLQ WSNIGNSPTE GDEFSFLLSM QMMLLDAAVY GLLAWYLDQV FPGDYGTPLP WYFLLQESYW LGGEGCSTR EERALEKTEP LTEETEDPEH PEGIHDSFFE REHPGWVPGV CVKNLVKIFE PCGRPAVDRL NITFYENQIT A FLGHNGAG KTTTLSILTG LLPPTSGTVL VGGRDIETSL DAVRQSLGMC PQHNILFHHL TVAEHMLFYA QLKGKSQEEA QL EMEAMLE DTGLHHKRNE EAQDLSGGMQ RKLSVAIAFV GDAKVVILDE PTSGVDPYSR RSIWDLLLKY RSGRTIIMST HHM DEADLL GDRIAIIAQG RLYCSGTPLF LKNCFGTGLY LTLVRKMKNI QSQRKGSEGT CSCSSKGFST TCPAHVDDLT PEQV LDGDV NELMDVVLHH VPEAKLVECI GQELIFLLPN KNFKHRAYAS LFRELEETLA DLGLSSFGIS DTPLEEIFLK VTEDS DSGP LFAGGAQQKR ENVNPRHPCL GPREKAGQTP QDSNVCSPGA PAAHPEGQPP PEPECPGPQL NTGTQLVLQH VQALLV KRF QHTIRSHKDF LAQIVLPATF VFLALMLSIV IPPFGEYPAL TLHPWIYGQQ YTFFSMDEPG SEQFTVLADV LLNKPGF GN RCLKEGWLPE YPCGNSTPWK TPSVSPNITQ LFQKQKWTQV NPSPSCRCST REKLTMLPEC PEGAGGLPPP QRTQRSTE I LQDLTDRNIS DFLVKTYPAL IRSSLKSKFW VNEQRYGGIS IGGKLPVVPI TGEALVGFLS DLGRIMNVSG GPITREASK EIPDFLKHLE TEDNIKVWFN NKGWHALVSF LNVAHNAILR ASLPKDRSPE EYGITVISQP LNLTKEQLSE ITVLTTSVDA VVAICVIFS MSFVPASFVL YLIQERVNKS KHLQFISGVS PTTYWVTNFL WDIMNYSVSA GLVVGIFIGF QKKAYTSPEN L PALVALLL LYGWAVIPMM YPASFLFDVP STAYVALSCA NLFIGINSSA ITFILELFEN NRTLLRFNAV LRKLLIVFPH FC LGRGLID LALSQAVTDV YARFGEEHSA NPFHWDLIGK NLFAMVVEGV VYFLLTLLVQ RHFFLSQWIA EPTKEPIVDE DDD VAEERQ RIITGGNKTD ILRLHELTKI YPGTSSPAVD RLCVGVRPGE CFGLLGVNGA GKTTTFKMLT GDTTVTSGDA TVAG KSILT NISEVHQNMG YCPQFDAIDE LLTGREHLYL YARLRGVPAE EIEKVANWSI KSLGLTVYAD CLAGTYSGGN KRKLS TAIA LIGCPPLVLL DEPTTGMDPQ ARRMLWNVIV SIIREGRAVV LTSHSMEECE ALCTRLAIMV KGAFRCMGTI QHLKSK FGD GYIVTMKIKS PKDDLLPDLN PVEQFFQGNF PGSVQRERHY NMLQFQVSSS SLARIFQLLL SHKDSLLIEE YSVTQTT LD QVFVNFAKQQ TESHDLPLHP RAAGASRQAQ D

UniProtKB: Retinal-specific phospholipid-transporting ATPase ABCA4

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Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 1 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER

MacromoleculeName: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / type: ligand / ID: 5 / Number of copies: 2 / Formula: ANP
Molecular weightTheoretical: 506.196 Da
Chemical component information

ChemComp-ANP:
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / AMP-PNP, energy-carrying molecule analogue*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

BufferpH: 7.4
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 825604
Startup modelType of model: OTHER / Details: Ab initio reconstruction
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.95 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 149336
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION

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