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- EMDB-36695: Cryo-EM structure of rat megalin wingA -

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Basic information

Entry
Database: EMDB / ID: EMD-36695
TitleCryo-EM structure of rat megalin wingA
Map datapostprocess map
Sample
  • Complex: megalin dimer at pH7.4
    • Protein or peptide: LDL receptor related protein 2
    • Protein or peptide: unclear peptide
    • Protein or peptide: unclear peptide
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 2-acetamido-2-deoxy-alpha-D-galactopyranose
  • Ligand: CALCIUM ION
Keywordsendocytosis receptor / ENDOCYTOSIS
Function / homology
Function and homology information


endocytosis / calcium ion binding / membrane
Similarity search - Function
: / Complement Clr-like EGF domain / Complement Clr-like EGF-like / Low-density lipoprotein receptor repeat class B / LDL-receptor class B (LDLRB) repeat profile. / LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A, conserved site / LDL-receptor class A (LDLRA) domain signature. ...: / Complement Clr-like EGF domain / Complement Clr-like EGF-like / Low-density lipoprotein receptor repeat class B / LDL-receptor class B (LDLRB) repeat profile. / LDLR class B repeat / Low-density lipoprotein-receptor YWTD domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A, conserved site / LDL-receptor class A (LDLRA) domain signature. / LDL-receptor class A (LDLRA) domain profile. / : / Calcium-binding EGF domain / Low-density lipoprotein receptor domain class A / Low-density lipoprotein (LDL) receptor class A repeat / LDL receptor-like superfamily / Six-bladed beta-propeller, TolB-like / Coagulation Factor Xa inhibitory site / EGF-type aspartate/asparagine hydroxylation site / EGF-like calcium-binding, conserved site / Calcium-binding EGF-like domain signature. / Aspartic acid and asparagine hydroxylation site. / EGF-like calcium-binding domain / Calcium-binding EGF-like domain / Epidermal growth factor-like domain. / EGF-like domain profile. / Growth factor receptor cysteine-rich domain superfamily / EGF-like domain signature 2. / EGF-like domain signature 1. / EGF-like domain / Prokaryotic membrane lipoprotein lipid attachment site profile.
Similarity search - Domain/homology
LDL receptor related protein 2
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsGoto S / Tsutsumi A / Lee Y / Hosojima M / Kabasawa H / Komochi K / Yun-san L / Nagatoshi S / Tsumoto K / Nishizawa T ...Goto S / Tsutsumi A / Lee Y / Hosojima M / Kabasawa H / Komochi K / Yun-san L / Nagatoshi S / Tsumoto K / Nishizawa T / Kikkawa M / Saito A
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS) Japan
CitationJournal: Proc Natl Acad Sci U S A / Year: 2024
Title: Cryo-EM structures elucidate the multiligand receptor nature of megalin.
Authors: Sawako Goto / Akihisa Tsutsumi / Yongchan Lee / Michihiro Hosojima / Hideyuki Kabasawa / Koichi Komochi / Satoru Nagatoishi / Kazuya Takemoto / Kouhei Tsumoto / Tomohiro Nishizawa / Masahide ...Authors: Sawako Goto / Akihisa Tsutsumi / Yongchan Lee / Michihiro Hosojima / Hideyuki Kabasawa / Koichi Komochi / Satoru Nagatoishi / Kazuya Takemoto / Kouhei Tsumoto / Tomohiro Nishizawa / Masahide Kikkawa / Akihiko Saito /
Abstract: Megalin (low-density lipoprotein receptor-related protein 2) is a giant glycoprotein of about 600 kDa, mediating the endocytosis of more than 60 ligands, including those of proteins, peptides, and ...Megalin (low-density lipoprotein receptor-related protein 2) is a giant glycoprotein of about 600 kDa, mediating the endocytosis of more than 60 ligands, including those of proteins, peptides, and drug compounds [S. Goto, M. Hosojima, H. Kabasawa, A. Saito, , 106393 (2023)]. It is expressed predominantly in renal proximal tubule epithelial cells, as well as in the brain, lungs, eyes, inner ear, thyroid gland, and placenta. Megalin is also known to mediate the endocytosis of toxic compounds, particularly those that cause renal and hearing disorders [Y. Hori , , 1783-1791 (2017)]. Genetic megalin deficiency causes Donnai-Barrow syndrome/facio-oculo-acoustico-renal syndrome in humans. However, it is not known how megalin interacts with such a wide variety of ligands and plays pathological roles in various organs. In this study, we elucidated the dimeric architecture of megalin, purified from rat kidneys, using cryoelectron microscopy. The maps revealed the densities of endogenous ligands bound to various regions throughout the dimer, elucidating the multiligand receptor nature of megalin. We also determined the structure of megalin in complex with receptor-associated protein, a molecular chaperone for megalin. The results will facilitate further studies on the pathophysiology of megalin-dependent multiligand endocytic pathways in multiple organs and will also be useful for the development of megalin-targeted drugs for renal and hearing disorders, Alzheimer's disease [B. V. Zlokovic , , 4229-4234 (1996)], and other illnesses.
History
DepositionJun 30, 2023-
Header (metadata) releaseJun 19, 2024-
Map releaseJun 19, 2024-
UpdateJun 19, 2024-
Current statusJun 19, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36695.png

Downloads & links

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Map

FileDownload / File: emd_36695.map.gz / Format: CCP4 / Size: 67 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationpostprocess map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.41 Å/pix.
x 260 pix.
= 366.86 Å		1.41 Å/pix.
x 260 pix.
= 366.86 Å		1.41 Å/pix.
x 260 pix.
= 366.86 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.411 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.13691162 - 0.2574692
Average (Standard dev.)0.0001958196 (±0.0036216588)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions260260260
Spacing260260260
CellA=B=C: 366.86002 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_36695_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: halfmap2

Fileemd_36695_half_map_1.map
Annotationhalfmap2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: halfmap1

Fileemd_36695_half_map_2.map
Annotationhalfmap1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : megalin dimer at pH7.4

EntireName: megalin dimer at pH7.4
Components
  • Complex: megalin dimer at pH7.4
    • Protein or peptide: LDL receptor related protein 2
    • Protein or peptide: unclear peptide
    • Protein or peptide: unclear peptide
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 2-acetamido-2-deoxy-alpha-D-galactopyranose
  • Ligand: CALCIUM ION

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Supramolecule #1: megalin dimer at pH7.4

SupramoleculeName: megalin dimer at pH7.4 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 1.2 MDa

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Macromolecule #1: LDL receptor related protein 2

MacromoleculeName: LDL receptor related protein 2 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 519.871438 KDa
SequenceString: MERGAAAAAW MLLLAIAACL EPVSSQECGS GNFRCDNGYC IPASWRCDGT RDCLDDTDEI GCPPRSCESG LFLCPAEGTC IPSSWVCDE DKDCSDGADE QQNCAGTTCS AQQMTCSNGQ CIPSEYRCDH VSDCPDGSDE RNCHYPTCDQ LTCANGACYN T SQRCDQKV ...String:
MERGAAAAAW MLLLAIAACL EPVSSQECGS GNFRCDNGYC IPASWRCDGT RDCLDDTDEI GCPPRSCESG LFLCPAEGTC IPSSWVCDE DKDCSDGADE QQNCAGTTCS AQQMTCSNGQ CIPSEYRCDH VSDCPDGSDE RNCHYPTCDQ LTCANGACYN T SQRCDQKV DCRDSSDEAN CTTLCSQKEF ECGSGECILR AYVCDHDNDC EDNSDERNCN YDTCGGHQFT CSNGQCINQN WV CDGDDDC QDSGDEDGCE SNQSHHRCYP REWACPGSGR CISIDKVCDG VPDCPEGDDE NNVTSGRTCG MGVCSVLNCE YQC HQTPFG GECFCPPGHI INSNDSRTCI DFDDCQIWGI CDQKCENRQG RHQCLCEEGY ILERGQHCKS SDSFSAASVI FSNG RDLLV GDLHGRNFRI LAESKNRGMV MGVDFHYQKH RVFWTDPMQE KVFSTDINGL NTQEILNVSV DTPENLAVDW INNKL YLVE TKVNRIDVVN LEGNQRVTLI TENLGHPRGI ALDPTVGYLF FSDWGSLSGQ PKVERAFMDG SNRKDLVTTK VGWPAG ITL DLVSKRVYWV DSRYDYIETV TYDGIQRKTV ARGGSLVPHP FGISLFEEHV FFTDWTKMAV MKASKFTETN PQVYHQS SL RPHGVTVYHA LRQPNATNPC GSNNGGCAQV CVLSHRTDNG GLGYRCKCEF GFELDDDEHR CVAVKNFLLF SSKTAVRG I PFTLSTQEDV MVPVTGSPSF FVGIDFDAQH STVFYSDLSK DIIYKQKIDG TGKEVITANR LESVECLTFD WISRNLYWT DGGLKSVTVL RLADKSRRQI ISNLNNPRSI VVHPTAGYMF LSDWFRPAKI MRAWSDGSHL MPIVNTSLGW PNGLAIDWSA SRLYWVDAF FDKIEHSTLD GLDRKRLGHV DQMTHPFGLT VFKDNVFITD WRLGAIIRVR KSDGGDMTVI RRGISSVMHV K AYDADLQT GSNYCSQTTH ANGDCSHFCF PVPNFQRVCG CPYGMKLQRD QMTCEGDPAR EPPTQQCGSL SFPCNNGKCV PS FFRCDGV DDCHDNSDEH QCGVFNNTCS PSAFACVRGG QCIPGQWHCD RQNDCLDGSD EQNCPTHATS STCPSTSFTC DNH VCIPKD WVCDTDNDCS DGSDEKNCQA SGTCQPTQFR CPDHRCISPL YVCDGDKDCA DGSDEAGCVL NCTSAQFKCA DGSS CINSR YRCDGVYDCR DNSDEAGCPT RPPGMCHLDE FQCQGDGTCI PNTWECDGHP DCIHGSDEHT GCVPKTCSPT HFLCD NGNC IYKAWICDGD NDCRDMSDEK DCPTQPFHCP STQWQCPGYS TCINLSALCD GVFDCPNGTD ESPLCNQDSC SHFNGG CTH QCMQGPFGAT CLCPLGYQLA NDTKTCEDIN ECDIPGFCSQ HCVNMRGSFR CACDPEYTLE SDGRTCKVTG SENPLLV VA SRDKIIVDNI TAHTHNLYSL VQDVSFVVAL DFDSVTGRVF WSDLLQGKTW SVFQNGTDKR VVHDSGLSVT EMIAVDWI G RNLYWTDYAL ETIEVSKIDG SHRTVLISKN VTKPRGLALD PRMGDNVMFW SDWGHHPRIE RASMDGTMRT VIVQEKIYW PCGLSIDYPN RLIYFMDAYL DYIEFCDYDG HNRRQVIASD LVLHHPHALT LFEDFVYWTD RGTRQVMQAN KWHGGNQSVV MYSVHQPLG ITAIHPSRQP PSRNPCASAS CSHLCLLSAQ APRHYSCACP SGWNLSDDSV NCVRGDQPFL MSVRDNIIFG I SLDPEVKS NDAMVPISGI QHGYDVEFDD SEQFIYWVEN PGEIHRVKTD GSNRTVFAPL SLLGSSLGLA LDWVSRNIYY TT PASRSIE VLTLKGDTRY GKTLIANDGT PLGVGFPVGI AVDPARGKLY WSDHGTDSGV PAKIASANMD GTSLKILFTG NLQ HLEVVT LDIQEQKLYW AVTSRGVIER GNVDGTERMI LVHHLAHPWG LVVYGSFLYY SDEQYEVIER VDKSSGNNKV VLRD NVPYL RGLRVYHRRN AADSSNGCSN NPNACQQICL PVPGGMFSCA CASGFKLSPD GRSCSPYNSF MVVSMLPAVR GFSLE LSDH SEAMVPVAGQ GRNVLHADVD VANGFIYWCD FSSSVRSSNG IRRIKPDGSN FTNVVTYGIG ANGIRGVALD WAAGNL YFT NAFVYETLIE VLRINTTYRR VLLKVSVDMP RHIIVDPKHR YLFWADYGQK PKIERSFLDC TNRTVLVSEG IVTPRGL AM DHDTGYIYWV DDSLDLIARI HLDGGESQVV RYGSRYPTPY GITVFGESII WVDRNLKKVF QASKQPGNTD PPVVIRDK I NLLRDVTIFD EHAQPLSPAE LNNNPCLQSN GGCSHFCFAL PELPTPRCGC AFGTLGNDGK SCATSQEDFL IYSLNNSLR SLHFDPRDHS LPFQVISVAG TAIALDYDRR NNRIFFTQKL NSLRGQISYV SLYSGSSSPT VLLSNIGVTD GIAFDWINRR IYYSDFSNQ TINSMAEDGS NRAVIARVSK PRAIVLDPCR GYMYWTDWGT NAKIERATLG GNFRVPIVNT SLVWPNGLAL D LETDLLYW ADASLQKIER STLTGTNREV VVSTAFHSFG LTVYGQYIYW TDLYTRKIYR ANKYDGSDLV AMTTRLPTQP SG ISTVVKT QRQQCSNPCD QFNGGCSHIC APGPNGAECQ CPHEGNWYLA NDNKYCVVDT GTRCNQLQFT CLNGHCINQD WKC DNDNDC GDGSDELPTV CAFHTCRSTA FTCGNGRCVP YHYRCDYYND CGDNSDEAGC LFRNCNSTTE FTCSNGRCIP LSYV CNGIN NCHDNDTSDE KNCPPHTCPP DFTKCQTTNI CVPRAFLCDG DNDCGDGSDE NPIYCASHTC RSNEFQCLSP QRCIP SYWF CDGEADCADG SDEPDTCGHS VNTCRASQFQ CDNGRCISGN WVCDGDNDCG DMSDEDQRHH CELQNCSSTQ FTCVNS RPP NRRCIPQYWV CDGDADCSDA LDELQNCTMR TCSAGEFSCA NGRCVRQSFR CDRRNDCGDY SDERGCSYPP CHANQFT CQ NGRCIPRFFV CDEDNDCGDG SDEQEHLCHT PEPTCPLHQF RCDNGHCIEM GRVCNHVDDC SDNSDEKGCG INECLDSS I SRCDHNCTDT ITSFYCSCLP GYKLMSDKRS CVDIDECKES PQLCSQKCEN VVGSYICKCA PGYIREPDGK SCRQNSNIE PYLIFSNRYY IRNLTTDGSS YSLILQGLGN VVALDFDRVE KRLYWIDAEK QIIERMFLNK TNRETIINHR LRRAESLAVD WVSRKLYWL DAILDCLFVS DLEGRHRKMI AQHCVDANNT FCFEHPRGIV LHPQRGHVYW ADWGVHAYIG RIGMDGTNKS V IISTKIEW PNAITIDYTN DLLYWADAHL GYIEFSDLEG HHRHTVYDGS LPHPFALTIF EDTVFWTDWN TRTVEKGNKY DG SGRVVLV NTTHKPFDIH VYHPYRQPIM SNPCGTNNGG CSHLCLIKAG GRGFTCACPD DFQTVQLRDR TLCMPMCSST QFL CGNNEK CIPIWWKCDG QKDCSDGSDE PDLCPHRFCR LGQFQCRDGN CTSPQALCNA RQDCADGSDE DRVLCEHHRC ESNE WQCAN KRCIPQSWQC DSVNDCLDNS DEDTSHCASR TCRPGQFKCN NGRCIPQSWK CDVDNDCGDY SDEPIDECTT AAYNC DNHT EFSCKTNYRC IPQWAVCNGF DDCRDNSDEQ GCESVPCHPS GDFRCANHHC IPLRWKCDGT DDCGDNSDEE NCVPRE CSE SEFRCADQQC IPSRWVCDQE NDCGDNSDER DCEMKTCHPE HFQCTSGHCV PKALACDGRA DCLDASDESA CPTRFPN GT YCPAAMFECK NHVCIQSFWI CDGENDCVDG SDEEIHLCFN IPCESPQRFR CDNSRCVYGH QLCNGVDDCG DGSDEKEE H CRKPTHKPCT DTEYKCSNGN CISQHYVCDN VNDCGDLSDE TGCNLGDNRT CAENICEQNC TQLSSGGFIC SCRPGFKPS TLDKNSCQDI NECEEFGICP QSCRNSKGSY ECFCVDGFKS MSTHYGERCA ADGSPPLLLL PENVRIRKYN TSSEKFSEYL EEEEHIQTI DYDWDPEHIG LSVVYYTVLA QGSQFGAIKR AYIPNFESGS NNPIREVDLG LKYLMQPDGL AVDWVGRHIY W SDAKSQRI EVATLDGRYR KWLITTQLDQ PAAIAVNPKL GLMFWTDQGK QPKIESAWMN GEHRSVLVSE NLGWPNGLSI DY LNDDRVY WSDSKEDVIE AIKYDGTDRR LIINEAMKPF SLDIFEDKLY WVAKEKGEVW RQNKFGKENK EKVLVVNPWL TQV RIFHQL RYNQSVSNPC KQVCSHLCLL RPGGYSCACP QGSDFVTGST VQCDAASELP VTMPPPCRCM HGGNCYFDEN ELPK CKCSS GYSGEYCEVG LSRGIPPGTT MAVLLTFVIV IIVGALVLVG LFHYRKTGSL LPTLPKLPSL SSLAKPSENG NGVTF RSGA DVNMDIGVSP FGPETIIDRS MAMNEHFVME VGKQPVIFEN PMYAAKDNTS KVALAVQGPS TGAQVTVPEN VENQNY GRP IDPSEIVPEP KPASPGADEI QGKKWNIFKR KPKQTTNFEN PIYAEMDSEV KDAVAVAPPP SPSLPAKASK RNLTPGY TA TEDTFKDTAN LVKEDSDV

UniProtKB: LDL receptor related protein 2

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Macromolecule #2: unclear peptide

MacromoleculeName: unclear peptide / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 528.644 Da
SequenceString:
(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)

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Macromolecule #3: unclear peptide

MacromoleculeName: unclear peptide / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Molecular weightTheoretical: 472.537 Da
SequenceString:
(UNK)N(UNK)(UNK)(UNK)

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Macromolecule #6: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 6 / Number of copies: 5 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

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

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Macromolecule #7: 2-acetamido-2-deoxy-alpha-D-galactopyranose

MacromoleculeName: 2-acetamido-2-deoxy-alpha-D-galactopyranose / type: ligand / ID: 7 / Number of copies: 8 / Formula: A2G
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-A2G:
2-acetamido-2-deoxy-alpha-D-galactopyranose

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Macromolecule #8: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 8 / Number of copies: 18 / Formula: CA
Molecular weightTheoretical: 40.078 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

Concentration2.4 mg/mL
BufferpH: 7.4
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: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1.4) / Number images used: 101096
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.4)
Final 3D classificationSoftware - Name: RELION (ver. 3.1.4)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8jxb:
Cryo-EM structure of rat megalin wingAc

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