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- EMDB-61327: Structure of Engineered Coagulation Factor VIII with Enhanced Sec... -

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Basic information

Entry
Database: EMDB / ID: EMD-61327
TitleStructure of Engineered Coagulation Factor VIII with Enhanced Secretion and Coagulation Potential
Map dataunsharpened map
Sample
  • Complex: Engineered Coagulation Factor VIII with Enhanced Secretion and Coagulation Potential
    • Protein or peptide: Coagulation factor VIII
Keywordshemophilia / BLOOD CLOTTING
Function / homology
Function and homology information


Defective F8 accelerates dissociation of the A2 domain / Defective F8 binding to the cell membrane / Defective F8 secretion / Defective F8 sulfation at Y1699 / Gamma carboxylation, hypusinylation, hydroxylation, and arylsulfatase activation / Defective F8 binding to von Willebrand factor / blood coagulation, intrinsic pathway / Cargo concentration in the ER / COPII-coated ER to Golgi transport vesicle / Defective factor IX causes thrombophilia ...Defective F8 accelerates dissociation of the A2 domain / Defective F8 binding to the cell membrane / Defective F8 secretion / Defective F8 sulfation at Y1699 / Gamma carboxylation, hypusinylation, hydroxylation, and arylsulfatase activation / Defective F8 binding to von Willebrand factor / blood coagulation, intrinsic pathway / Cargo concentration in the ER / COPII-coated ER to Golgi transport vesicle / Defective factor IX causes thrombophilia / Defective cofactor function of FVIIIa variant / Defective F9 variant does not activate FX / COPII-mediated vesicle transport / Defective F8 cleavage by thrombin / : / : / endoplasmic reticulum-Golgi intermediate compartment membrane / platelet alpha granule lumen / acute-phase response / Golgi lumen / blood coagulation / Platelet degranulation / oxidoreductase activity / endoplasmic reticulum lumen / copper ion binding / : / extracellular region / plasma membrane
Similarity search - Function
Coagulation factor 5/8-like / : / Coagulation factors 5/8 type C domain (FA58C) signature 2. / Multicopper oxidases, conserved site / Multicopper oxidases signature 1. / Coagulation factors 5/8 type C domain (FA58C) signature 1. / Coagulation factor 5/8 C-terminal domain, discoidin domain / Coagulation factors 5/8 type C domain (FA58C) profile. / Multicopper oxidase, C-terminal / Multicopper oxidase ...Coagulation factor 5/8-like / : / Coagulation factors 5/8 type C domain (FA58C) signature 2. / Multicopper oxidases, conserved site / Multicopper oxidases signature 1. / Coagulation factors 5/8 type C domain (FA58C) signature 1. / Coagulation factor 5/8 C-terminal domain, discoidin domain / Coagulation factors 5/8 type C domain (FA58C) profile. / Multicopper oxidase, C-terminal / Multicopper oxidase / F5/8 type C domain / Coagulation factor 5/8 C-terminal domain / Multicopper oxidase, N-terminal / Multicopper oxidase / Cupredoxin / Galactose-binding-like domain superfamily
Similarity search - Domain/homology
Coagulation factor VIII
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.17 Å
AuthorsKio H / Osamu N / Tsukasa O
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Agency for Medical Research and Development (AMED) Japan
CitationJournal: Blood / Year: 2026
Title: Engineered coagulation factor VIII with enhanced secretion and coagulation potential for hemophilia A gene therapy.
Authors: Yuji Kashiwakura / Yuto Nakajima / Kio Horinaka / Tiago J S Lopes / Yuma Furuta / Yuki Yamaguchi / Nemekhbayar Baatartsogt / Morisada Hayakawa / Yuko Katakai / Susumu Uchiyama / Osamu Nureki ...Authors: Yuji Kashiwakura / Yuto Nakajima / Kio Horinaka / Tiago J S Lopes / Yuma Furuta / Yuki Yamaguchi / Nemekhbayar Baatartsogt / Morisada Hayakawa / Yuko Katakai / Susumu Uchiyama / Osamu Nureki / Keiji Nogami / Tsukasa Ohmori /
Abstract: The major challenges of gene therapy for hemophilia A using adeno-associated virus (AAV) vectors are reducing vector doses and the long-term maintenance of stable factor VIII (FVIII). In this study, ...The major challenges of gene therapy for hemophilia A using adeno-associated virus (AAV) vectors are reducing vector doses and the long-term maintenance of stable factor VIII (FVIII). In this study, we developed engineered human B-domain-deleted FVIIIs (FVIIISQ) with enhanced secretion and coagulation potential. Intracellular accumulation was markedly reduced in some engineered FVIIISQ, resulting in reduced unfolded protein responses. The administration of AAV vectors carrying engineered FVIIISQ to hemophilia A mice resulted in ∼8-fold higher FVIII activity and 4-fold higher FVIII antigen levels compared with wild-type FVIIISQ administration. The specific FVIII activity of the engineered FVIIISQ was 3.6 times higher than that of the wild-type FVIIISQ, and its binding to activated coagulation factor IX was significantly enhanced, which is supported by the structural analysis. In macaques, the administration of AAV5 vector carrying the engineered FVIIISQ without CpG sequences resulted in a supraphysiological increase in plasma FVIII activity at a dose one-thirtieth that of valoctocogene roxaparvovec (2 × 1012 vector genome per kg). The engineered FVIIISQ may thus provide stable, long-term therapeutic efficacy in AAV-mediated hemophilia A gene therapy even at low doses.
History
DepositionAug 27, 2024-
Header (metadata) releaseSep 10, 2025-
Map releaseSep 10, 2025-
UpdateMar 25, 2026-
Current statusMar 25, 2026Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_61327.png

Downloads & links

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Map

FileDownload / File: emd_61327.map.gz / Format: CCP4 / Size: 14.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationunsharpened map
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.11 Å/pix.
x 156 pix.
= 172.645 Å		1.11 Å/pix.
x 156 pix.
= 172.645 Å		1.11 Å/pix.
x 156 pix.
= 172.645 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1067 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.035
Minimum - Maximum-0.10909623 - 0.29203698
Average (Standard dev.)-0.001702833 (±0.011236227)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin343434
Dimensions156156156
Spacing156156156
CellA=B=C: 172.64519 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_61327_msk_1.map
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Half map: #1

Fileemd_61327_half_map_1.map
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Half map: #2

Fileemd_61327_half_map_2.map
Projections & Slices
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Sample components

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Entire : Engineered Coagulation Factor VIII with Enhanced Secretion and Co...

EntireName: Engineered Coagulation Factor VIII with Enhanced Secretion and Coagulation Potential
Components
  • Complex: Engineered Coagulation Factor VIII with Enhanced Secretion and Coagulation Potential
    • Protein or peptide: Coagulation factor VIII

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Supramolecule #1: Engineered Coagulation Factor VIII with Enhanced Secretion and Co...

SupramoleculeName: Engineered Coagulation Factor VIII with Enhanced Secretion and Coagulation Potential
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Coagulation factor VIII

MacromoleculeName: Coagulation factor VIII / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 167.452469 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MQIELSTCFF LCLLPFCFSA TRRYYLGAVE LSWDYMQSDL GELHVDTRFP PRVPKSFPFN TSVVYKKTLF VEFTDHLFNI AKPRPPWMG LLGPTIQAEV YDTVVITLKN MASHPVSLHA VGVSYWKASE GAEYDDQTSQ REKEDDKVFP GGSHTYVWQV L KENGPMAS ...String:
MQIELSTCFF LCLLPFCFSA TRRYYLGAVE LSWDYMQSDL GELHVDTRFP PRVPKSFPFN TSVVYKKTLF VEFTDHLFNI AKPRPPWMG LLGPTIQAEV YDTVVITLKN MASHPVSLHA VGVSYWKASE GAEYDDQTSQ REKEDDKVFP GGSHTYVWQV L KENGPMAS DPPCLTYSYL SHVDLVKDLN SGLIGALLVC REGSLAKEKT QTLHKFILLF AVFDEGKSWH SETNNSLTQD RD AASARAQ PKMHTVNGYV NRSLPGLIGC HRKSVYWHVI GMGTTPEVHS IFLEGHTFLV RNHRQASLEI SPITFLTAQT LLM DLGQFL LFCHISSHQH DGMEAYVKVD SCPEEPQLRM KNNEEAEDYD DDLTDSEMDV VRFDDDNSPP FIQIRSVAKK HPKT WVHYI AAEEEDWDYA PLVLAPDDRS YKSLYLNNGP QRIGRKYKKV RFMAYTDETF KTREAIQHES GILGPLLYGE VGDTL LIIF KNQASRPYNI YPHGITDVRP LHSGRLPKGV KHLKDMPILP GEIFKYKWTV TVEDGPTKSD PRCLTRYYSS FVNLER DLA SGLIGPLLIC YKESVDQRGN QMMSDKRNVI LFSVFDENRS WYLTENIQRF LPNPAGVQPE DPEFQASNIM HSINGYV FD SLQLSVCLHE VAYWYILSVG AQTDFLSVFF SGYTFKHKMV YEDTLTLFPF SGETVFMSME NPGLWILGCH NSDFRNRG M TALLKVSSCD KNTGDYYEDS YEDIPAYLLS KNNVIEPRSF SQNPPVLKRH QREITRTTLQ PDEEKIDYDD TFSVEMKKE DFDIYGEDEN QSPRSFQKKT RHYFIAAVER LWDYGMSSSP HVLRNRAQSG SVPQFKKVVF QEFTDGSFTQ PLYRGELNEH LGLLGPYIR AEVEDNIMVT FKNQASRPYS FYSSLISYEE DQRQGAEPRK NFVKPNETKT YFWKVQHHMA PTKDEFDCKA W AYFSDVDL EKDVHSGLIG PLLVCRTNTL NPAHGRQVTV QEFALFFTIF DETKSWYFTE NMERNCRAPC NIQMEDPTFK EN YRFHAIN GYIMDTLPGL VMAQDQRIRW YLLSMGSNEN IHSIHFSGHV FTVRKKEEYK MALYNLYPGV FETVEMLPSK VGI WRVECL IGEHLQAGMS TLFLVYSKKC QTPLGMASGH IRDFQITASG QYGQWAPKLA RLHYSGSINA WSTKEPFSWI KVDL LAPMI IHGIMTQGAR QKFSSLYISQ FIIMYSLDGK KWQTYRGNST GTLMVFFGNV DSSGIKHNIF NPPIIARYIR LHPTH YSIR STLRMELMGC DLNSCSMPLG MESKAISDAQ ITASSYLTNM FATWSPSQAR LHLQGRSNAW RPQVNNPKEW LQVDFQ KTM KVTGVTTQGV KSLLTSMYVK EFLISSSQDG HQWTLFLQNG KVKVFQGNQD SFTPVVNALD PPLLTRYLRI HPQSWAH HI ALRLEVLGCE AQDLY

UniProtKB: Coagulation factor VIII, Coagulation factor VIII

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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: 7.3
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (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: 1.6 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.17 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 218890
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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