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- PDB-5ywo: Structure of JEV-2F2 Fab complex -

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Basic information

Entry
Database: PDB / ID: 5ywo
TitleStructure of JEV-2F2 Fab complex
Components
  • 2F2 heavy chain
  • 2F2 light chain
  • JEV E protein
  • JEV M protein
KeywordsVIRUS / Flaviviruses / Viral encephalitis / neutralizing monoclonal antibodies / structural analysis
Function / homology
Function and homology information


symbiont-mediated suppression of host JAK-STAT cascade via inhibition of host TYK2 activity / flavivirin / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT2 activity / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / viral capsid / nucleoside-triphosphate phosphatase / double-stranded RNA binding / mRNA (guanine-N7)-methyltransferase / methyltransferase cap1 / clathrin-dependent endocytosis of virus by host cell ...symbiont-mediated suppression of host JAK-STAT cascade via inhibition of host TYK2 activity / flavivirin / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT2 activity / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / viral capsid / nucleoside-triphosphate phosphatase / double-stranded RNA binding / mRNA (guanine-N7)-methyltransferase / methyltransferase cap1 / clathrin-dependent endocytosis of virus by host cell / mRNA (nucleoside-2'-O-)-methyltransferase activity / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / host cell surface / RNA helicase activity / host cell endoplasmic reticulum membrane / host cell perinuclear region of cytoplasm / protein dimerization activity / RNA helicase / induction by virus of host autophagy / RNA-directed RNA polymerase / viral RNA genome replication / RNA-dependent RNA polymerase activity / serine-type endopeptidase activity / fusion of virus membrane with host endosome membrane / viral envelope / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / host cell nucleus / virion attachment to host cell / virion membrane / structural molecule activity / ATP hydrolysis activity / proteolysis / extracellular region / ATP binding / membrane / metal ion binding
Similarity search - Function
: / Flavivirus envelope glycoprotein E, stem/anchor domain / RNA-directed RNA polymerase, thumb domain, Flavivirus / Flavivirus RNA-directed RNA polymerase, thumb domain / Flavivirus capsid protein C superfamily / : / Flavivirus non-structural protein NS2B / Flavivirus NS3 helicase, C-terminal helical domain / Genome polyprotein, Flavivirus / Flavivirus non-structural protein NS4A ...: / Flavivirus envelope glycoprotein E, stem/anchor domain / RNA-directed RNA polymerase, thumb domain, Flavivirus / Flavivirus RNA-directed RNA polymerase, thumb domain / Flavivirus capsid protein C superfamily / : / Flavivirus non-structural protein NS2B / Flavivirus NS3 helicase, C-terminal helical domain / Genome polyprotein, Flavivirus / Flavivirus non-structural protein NS4A / Flavivirus non-structural protein NS2B / Flavivirus capsid protein C / Flavivirus non-structural protein NS4B / mRNA cap 0/1 methyltransferase / Flavivirus capsid protein C / Flavivirus non-structural protein NS4B / Flavivirus non-structural protein NS4A / Flavivirus NS2B domain profile. / mRNA cap 0 and cap 1 methyltransferase (EC 2.1.1.56 and EC 2.1.1.57) domain profile. / Flavivirus non-structural protein NS2A / Flavivirus non-structural protein NS2A / Flavivirus NS3, petidase S7 / Peptidase S7, Flavivirus NS3 serine protease / Flavivirus NS3 protease (NS3pro) domain profile. / Envelope glycoprotein M, flavivirus / Flavivirus envelope glycoprotein M / RNA-directed RNA polymerase, flavivirus / Flavivirus RNA-directed RNA polymerase, fingers and palm domains / Flavivirus non-structural Protein NS1 / Flavivirus non-structural protein NS1 / Envelope glycoprotein M superfamily, flavivirus / Flavivirus polyprotein propeptide / Flavivirus polyprotein propeptide superfamily / Flavivirus polyprotein propeptide / Flaviviral glycoprotein E, central domain, subdomain 1 / Flaviviral glycoprotein E, central domain, subdomain 2 / Flavivirus envelope glycoprotein E, Stem/Anchor domain / Flavivirus glycoprotein E, immunoglobulin-like domain / Flavivirus envelope glycoprotein E, Stem/Anchor domain superfamily / Flavivirus glycoprotein, immunoglobulin-like domain / Flavivirus glycoprotein central and dimerisation domain / Flavivirus glycoprotein, central and dimerisation domains / Ribosomal RNA methyltransferase, FtsJ domain / FtsJ-like methyltransferase / Flavivirus/Alphavirus glycoprotein, immunoglobulin-like domain superfamily / Flavivirus glycoprotein, central and dimerisation domain superfamily / Flaviviral glycoprotein E, dimerisation domain / DEAD box, Flavivirus / Flavivirus DEAD domain / helicase superfamily c-terminal domain / Immunoglobulin E-set / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / RNA-directed RNA polymerase, catalytic domain / RdRp of positive ssRNA viruses catalytic domain profile. / S-adenosyl-L-methionine-dependent methyltransferase superfamily / Peptidase S1, PA clan / DNA/RNA polymerase superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Genome polyprotein / Genome polyprotein
Similarity search - Component
Biological speciesJapanese encephalitis virus
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.7 Å
AuthorsQiu, X. / Lei, Y.F. / Yang, P. / Gao, Q. / Wang, N. / Cao, L. / Yuan, S. / Wang, X. / Xu, Z.K. / Rao, Z.
Funding support China, 1items
OrganizationGrant numberCountry
National Key Research and Development Program of China2016YFC1200400 China
CitationJournal: Nat Microbiol / Year: 2018
Title: Structural basis for neutralization of Japanese encephalitis virus by two potent therapeutic antibodies.
Authors: Xiaodi Qiu / Yingfeng Lei / Pan Yang / Qiang Gao / Nan Wang / Lei Cao / Shuai Yuan / Xiaofang Huang / Yongqiang Deng / Wenyu Ma / Tianbing Ding / Fanglin Zhang / Xingan Wu / Junjie Hu / Shan- ...Authors: Xiaodi Qiu / Yingfeng Lei / Pan Yang / Qiang Gao / Nan Wang / Lei Cao / Shuai Yuan / Xiaofang Huang / Yongqiang Deng / Wenyu Ma / Tianbing Ding / Fanglin Zhang / Xingan Wu / Junjie Hu / Shan-Lu Liu / Chengfeng Qin / Xiangxi Wang / Zhikai Xu / Zihe Rao /
Abstract: Japanese encephalitis virus (JEV), closely related to dengue, Zika, yellow fever and West Nile viruses, remains neglected and not well characterized . JEV is the leading causative agent of ...Japanese encephalitis virus (JEV), closely related to dengue, Zika, yellow fever and West Nile viruses, remains neglected and not well characterized . JEV is the leading causative agent of encephalitis, and is responsible for thousands of deaths each year in Asia. Humoral immunity is essential for protecting against flavivirus infections and passive immunization has been demonstrated to be effective in curing disease. Here, we demonstrate that JEV-specific monoclonal antibodies, 2F2 and 2H4, block attachment of the virus to its receptor and also prevent fusion of the virus. Neutralization of JEV by these antibodies is exceptionally potent and confers clear therapeutic benefit in mouse models. A single 20 μg dose of these antibodies resulted in 100% survival and complete clearance of JEV from the brains of mice. The 4.7 Å and 4.6 Å resolution cryo-electron microscopy structures of JEV-2F2-Fab and JEV-2H4-Fab complexes, together with the crystal structure of 2H4 Fab and our recent near-atomic structure of JEV , unveil the nature and location of epitopes targeted by the antibodies. Both 2F2 and 2H4 Fabs bind quaternary epitopes that span across three adjacent envelope proteins. Our results provide a structural and molecular basis for the application of 2F2 and 2H4 to treat JEV infection.
History
DepositionNov 29, 2017Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 21, 2018Provider: repository / Type: Initial release
Revision 1.1Sep 12, 2018Group: Data collection / Source and taxonomy / Category: entity_src_nat
Item: _entity_src_nat.pdbx_ncbi_taxonomy_id / _entity_src_nat.pdbx_organism_scientific

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Structure visualization

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  • Biological unit as complete icosahedral assembly
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  • Biological unit as icosahedral pentamer
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  • Biological unit as icosahedral 23 hexamer
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  • Deposited structure unit
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Assembly

Deposited unit
A: JEV E protein
B: JEV M protein
C: JEV E protein
D: JEV M protein
E: JEV E protein
F: JEV M protein
M: 2F2 light chain
N: 2F2 heavy chain
Q: 2F2 light chain
R: 2F2 heavy chain


Theoretical massNumber of molelcules
Total (without water)280,09310
Polymers280,09310
Non-polymers00
Water0
1
A: JEV E protein
B: JEV M protein
C: JEV E protein
D: JEV M protein
E: JEV E protein
F: JEV M protein
M: 2F2 light chain
N: 2F2 heavy chain
Q: 2F2 light chain
R: 2F2 heavy chain
x 60


Theoretical massNumber of molelcules
Total (without water)16,805,584600
Polymers16,805,584600
Non-polymers00
Water0
TypeNameSymmetry operationNumber
point symmetry operation60
Buried area33300 Å2
ΔGint-170 kcal/mol
Surface area110050 Å2
2


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
point symmetry operation1
3
A: JEV E protein
B: JEV M protein
C: JEV E protein
D: JEV M protein
E: JEV E protein
F: JEV M protein
M: 2F2 light chain
N: 2F2 heavy chain
Q: 2F2 light chain
R: 2F2 heavy chain
x 5


  • icosahedral pentamer
  • 1.4 MDa, 50 polymers
Theoretical massNumber of molelcules
Total (without water)1,400,46550
Polymers1,400,46550
Non-polymers00
Water0
TypeNameSymmetry operationNumber
point symmetry operation5
4
A: JEV E protein
B: JEV M protein
C: JEV E protein
D: JEV M protein
E: JEV E protein
F: JEV M protein
M: 2F2 light chain
N: 2F2 heavy chain
Q: 2F2 light chain
R: 2F2 heavy chain
x 6


  • icosahedral 23 hexamer
  • 1.68 MDa, 60 polymers
Theoretical massNumber of molelcules
Total (without water)1,680,55860
Polymers1,680,55860
Non-polymers00
Water0
TypeNameSymmetry operationNumber
point symmetry operation6
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
6
A: JEV E protein
B: JEV M protein
C: JEV E protein
D: JEV M protein
E: JEV E protein
F: JEV M protein
M: 2F2 light chain
N: 2F2 heavy chain
Q: 2F2 light chain
R: 2F2 heavy chain
x 60


  • crystal asymmetric unit, crystal frame
  • 16.8 MDa, 600 polymers
Theoretical massNumber of molelcules
Total (without water)16,805,584600
Polymers16,805,584600
Non-polymers00
Water0
TypeNameSymmetry operationNumber
transform to crystal frame1
point symmetry operation60
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))
Noncrystallographic symmetry (NCS)NCS oper:
IDCodeMatrixVector
1given(1), (1), (1)
2generate(0.36180342, -0.26286551, -0.8944272), (0.58778526, 0.80901699, 6.0E-8), (0.72360678, -0.52573114, 0.44721358)872.6078, -192.84593, 172.48663
3generate(-0.67082035, 0.16245993, -0.72360683), (0.68819102, 0.49999999, -0.52573105), (0.27639317, -0.8506508, -0.44721363)1084.73608, 164.04445, 982.43502
4generate(-0.67082034, 0.68819102, 0.27639318), (0.16245994, 0.49999999, -0.8506508), (-0.72360682, -0.52573105, -0.44721363)343.23078, 577.46076, 1310.52402
5generate(0.36180342, 0.58778526, 0.72360679), (-0.2628655, 0.80901699, -0.52573115), (-0.89442719, 6.0E-8, 0.44721358)-327.17299, 476.07571, 703.34579
6generate(-0.86180345, -0.42532533, 0.27639316), (-0.42532532, 0.30901701, -0.85065084), (0.27639315, -0.85065084, -0.44721356)977.21752, 955.94214, 982.43501
7generate(-0.36180343, -0.2628655, 0.8944272), (-0.58778522, 0.80901701, -6.0E-8), (-0.7236068, -0.52573112, -0.44721358)354.89749, 378.48138, 1310.52402
8generate(0.3618034, -0.58778525, 0.7236068), (0.26286553, 0.809017, 0.52573112), (-0.89442719, -3.0E-8, 0.44721359)244.15428, -290.44023, 703.34583
9generate(0.30901699, -0.95105652, -5.0E-8), (0.95105651, 0.309017, 7.0E-8), (-5.0E-8, -7.0E-8, 1)798.03124, -126.39577, 6.0E-5
10generate(-0.44721364, -0.85065077, -0.27639328), (0.52573118, 1.0E-8, -0.85065077), (0.72360672, -0.52573119, 0.44721362)1251.08924, 643.9109, 172.48666
11generate(0.80901702, 0.58778523, 1.3E-7), (0.58778522, -0.80901702, 4.0E-8), (1.3E-7, 4.0E-8, -1)-192.84595, 593.51863, 971.9999
12generate(0.63819671, 0.2628655, -0.72360673), (-0.26286554, -0.809017, -0.52573112), (-0.7236067, 0.52573114, -0.4472137)399.75664, 1262.44023, 799.51338
13generate(-0.13819652, 0.42532534, -0.89442724), (-0.95105652, -0.30901699, -5.0E-8), (-0.27639323, 0.85065084, 0.44721351)781.14703, 1098.39575, -10.43497
14generate(-0.44721362, 0.85065078, -0.27639325), (-0.52573117, -1.0E-8, 0.85065078), (0.72360674, 0.52573116, 0.44721363)424.25667, 328.08908, -338.52405
15generate(0.13819654, 0.95105651, 0.27639326), (0.42532533, -0.30901702, 0.85065084), (0.89442723, 6.0E-8, -0.44721351)-177.7041, 16.05786, 268.65409
16generate(-0.94721356, -0.1624599, -0.27639329), (-0.1624599, -0.5, 0.8506508), (-0.27639329, 0.8506508, 0.44721356)1159.62845, 394.53923, -10.43494
17generate(-0.6381967, 0.26286551, 0.72360673), (0.26286551, -0.80901701, 0.52573111), (0.72360673, 0.52573112, 0.4472137)316.7381, 495.9243, -338.52406
18generate(0.44721346, -2.0E-8, 0.89442726), (-3.0E-8, -1, -2.0E-8), (0.89442725, -2.0E-8, -0.44721346)-166.03737, 972.00002, 268.65409
19generate(0.80901697, -0.58778528, 1.3E-7), (-0.58778528, -0.80901697, -4.0E-8), (1.3E-7, -4.0E-8, -1)378.48133, 1164.84592, 971.99994
20generate(-0.05278632, -0.68819101, -0.72360677), (-0.688191, -0.49999998, 0.52573108), (-0.72360676, 0.52573107, -0.4472137)1197.78787, 807.95552, 799.51344
21generate(-0.13819651, -0.42532534, -0.89442724), (0.95105651, -0.309017, 9.0E-8), (-0.27639326, -0.85065084, 0.44721351)1194.56326, 173.96875, 816.39766
22generate(-0.94721355, 0.16245992, -0.27639332), (0.16245992, -0.5, -0.8506508), (-0.27639332, -0.85065079, 0.44721355)1001.71743, 1063.46076, 816.39765
23generate(-0.44721363, 0.52573118, 0.72360673), (-0.85065076, -0.52573119), (-0.27639329, -0.85065076, 0.44721362)96.16761, 1154.92162, 816.39758
24generate(0.67082035, 0.16245991, 0.72360683), (-0.688191, 0.50000001, 0.52573105), (-0.2763932, -0.85065079, 0.44721363)-270.64712, 321.95554, 816.39755
25generate(0.86180344, -0.42532535, -0.27639316), (0.42532533, 0.309017, 0.85065085), (-0.27639319, -0.85065083, 0.44721356)408.19873, -284.30668, 816.3976
26generate(0.0527863, 0.68819099, 0.72360678), (-0.68819101, -0.5, 0.52573105), (0.72360675, -0.52573107, 0.4472137)-225.78785, 807.95555, 172.48655
27generate(0.94721356, 0.16245989, 0.27639332), (-0.16245994, -0.5, 0.85065079), (0.27639329, -0.8506508, -0.44721356)-187.62844, 394.53925, 982.43493
28generate(0.63819671, -0.26286554, -0.72360672), (0.26286549, -0.80901699, 0.52573115), (-0.72360672, -0.52573112, -0.4472137)655.26191, 495.92429, 1310.52405
29generate(-0.44721346, -0.89442726), (-1, 1.0E-8), (-0.89442725, 0.44721346)1138.0374, 971.99999, 703.3459
30generate(-0.80901699, 0.58778525, -1.3E-7), (-0.58778525, -0.809017, -5.0E-8), (-1.3E-7, 4.0E-8, 1)593.5187, 1164.84592, 5.0E-5
31generate(-0.3618034, 0.26286553, 0.8944272), (0.58778528, 0.80901698, 3.0E-8), (-0.72360678, 0.52573114, -0.44721358)99.3922, -192.84591, 799.51335
32generate(0.67082037, -0.16245992, 0.72360681), (0.68819099, 0.49999999, -0.52573108), (-0.27639316, 0.8506508, 0.44721363)-112.73608, 164.04448, -10.43503
33generate(0.67082035, -0.688191, -0.2763932), (0.1624599, 0.50000001, -0.85065079), (0.72360682, 0.52573105, 0.44721363)628.76923, 577.46076, -338.52403
34generate(-0.36180343, -0.58778523, -0.7236068), (-0.2628655, 0.80901701, -0.52573111), (0.89442719, -5.0E-8, -0.44721359)1299.173, 476.07568, 268.6542
35generate(-1, 3.0E-8), (4.0E-8, 1), (-1)972, -2.0E-5, 971.99998
36generate(0.4472136, -0.52573118, -0.72360675), (-0.85065078, 2.0E-8, -0.52573116), (0.27639328, 0.85065076, -0.44721363)875.83242, 1154.92161, 155.60241
37generate(-0.67082037, -0.16245989, -0.72360681), (-0.68819097, 0.50000001, 0.52573108), (0.2763932, 0.85065079, -0.44721362)1242.64712, 321.95551, 155.60243
38generate(-0.86180342, 0.42532536, 0.27639319), (0.42532536, 0.30901697, 0.85065084), (0.27639319, 0.85065084, -0.44721355)563.80126, -284.30668, 155.60238
39generate(0.13819654, 0.42532533, 0.89442724), (0.9510565, -0.30901702, 5.0E-8), (0.27639326, 0.85065083, -0.44721351)-222.56326, 173.96878, 155.60232
40generate(0.94721355, -0.16245993, 0.27639329), (0.16245988, -0.5, -0.8506508), (0.27639332, 0.85065079, -0.44721356)-29.7174, 1063.46078, 155.60234
41generate(-0.1381965, 0.95105652, -0.27639326), (-0.42532534, -0.309017, -0.85065084), (-0.89442723, 8.0E-8, 0.44721351)225.27716, 1256.30668, 703.34583
42generate(0.309017, 0.95105652, -5.0E-8), (-0.95105651, 0.30901699, -7.0E-8), (-5.0E-8, 7.0E-8, 1)-126.3957, 798.03125
43generate(0.67082037, 0.688191, -0.27639317), (-0.16245992, 0.49999999, 0.8506508), (0.7236068, -0.52573107, 0.44721363)-40.15244, -91.46076, 172.48656
44generate(0.44721365, 0.52573117, -0.72360673), (0.85065076, -2.0E-8, 0.52573119), (0.27639325, -0.85065077, -0.44721363)364.82168, -182.92162, 982.43497
45generate(-0.05278627, 0.688191, -0.72360678), (0.68819099, -0.50000002, -0.52573105), (-0.72360677, -0.52573105, -0.4472137)528.8662, 650.04447, 1310.52404
46generate(-0.36180339, 0.58778527, -0.72360679), (0.26286554, 0.80901698, 0.52573115), (0.89442719, 3.0E-8, -0.44721358)727.84571, -290.44024, 268.65415
47generate(-0.30901696, 0.95105653, 5.0E-8), (0.95105653, 0.30901696, 7.0E-8), (5.0E-8, 7.0E-8, -1)173.96875, -126.39576, 971.99993
48generate(0.44721365, 0.85065077, 0.27639326), (0.52573117, -2.0E-8, -0.85065078), (-0.72360672, 0.52573119, -0.44721363)-279.08922, 643.91093, 799.51333
49generate(0.86180344, 0.42532534, -0.27639319), (-0.42532535, 0.30901699, -0.85065084), (-0.27639315, 0.85065084, 0.44721356)-5.2175, 955.94216, -10.43502
50generate(0.36180341, 0.26286553, -0.8944272), (-0.58778524, 0.809017, -2.0E-8), (0.7236068, 0.52573112, 0.44721358)617.10252, 378.48138, -338.52404
51generate(0.4472136, -0.85065078, 0.27639328), (-0.52573118, 2.0E-8, 0.85065077), (-0.72360674, -0.52573116, -0.44721363)547.74334, 328.08908, 1310.52404
52generate(-0.13819653, -0.95105652, -0.27639324), (0.42532534, -0.30901698, 0.85065084), (-0.89442723, -6.0E-8, 0.44721351)1149.7041, 16.05783, 703.3459
53generate(-0.809017, -0.58778525, -1.3E-7), (0.58778525, -0.80901699, 4.0E-8), (-1.3E-7, -5.0E-8, 1)1164.84597, 593.5186, 9.0E-5
54generate(-0.63819672, -0.26286552, 0.72360671), (-0.26286552, -0.80901698, -0.52573115), (0.7236067, -0.52573114, 0.4472137)572.2434, 1262.44023, 172.48661
55generate(0.13819649, -0.42532535, 0.89442724), (-0.95105652, -0.30901697, -9.0E-8), (0.27639322, -0.85065084, -0.44721351)190.853, 1098.39576, 982.43496
56generate(0.05278629, -0.68819101, 0.72360676), (0.68819098, -0.5, -0.52573108), (0.72360678, 0.52573105, 0.4472137)443.13381, 650.04448, -338.52405
57generate(0.13819649, -0.95105653, 0.27639323), (-0.42532535, -0.30901697, -0.85065085), (0.89442723, -8.0E-8, -0.44721351)746.72287, 1256.30667, 268.65416
58generate(-0.30901703, -0.95105651, 5.0E-8), (-0.9510565, 0.30901703, -7.0E-8), (5.0E-8, -7.0E-8, -1)1098.39572, 798.03122, 972
59generate(-0.67082038, -0.68819098, 0.2763932), (-0.16245988, 0.50000001, 0.8506508), (-0.7236068, 0.52573107, -0.44721362)1012.15244, -91.46078, 799.51342
60generate(-0.44721363, -0.52573117, 0.72360674), (0.85065078, 0.52573116), (-0.27639325, 0.85065077, 0.44721363)607.17831, -182.92162, -10.43498

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Components

#1: Protein JEV E protein


Mass: 53508.684 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Japanese encephalitis virus / References: UniProt: P32886*PLUS
#2: Protein JEV M protein


Mass: 8250.488 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Japanese encephalitis virus / References: UniProt: P27395*PLUS
#3: Antibody 2F2 light chain


Mass: 23840.303 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#4: Antibody 2F2 heavy chain


Mass: 23567.473 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Complex of JEV with 2F2 Fab / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Japanese encephalitis virus / Strain: P3
Source (recombinant)Organism: Chlorocebus aethiops (grivet) / Cell: vero
Buffer solutionpH: 7.4 / Details: PBS
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 298 K / Details: blot for 3 seconds before plunging

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 1000 nm / Calibrated defocus max: 3000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Specimen holderSpecimen holder model: OTHER
Image recordingAverage exposure time: 8 sec. / Electron dose: 25 e/Å2 / Detector mode: OTHER / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 5 / Num. of real images: 1000
Image scansWidth: 4000 / Height: 4000

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Processing

SoftwareName: PHENIX / Version: 1.10.1_2155: / Classification: refinement
EM software
IDNameVersionCategory
4GctfCTF correction
7UCSF Chimeramodel fitting
10RELION1.4final Euler assignment
11RELIONclassification
12RELION1.43D reconstruction
13PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 14203
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionResolution: 4.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 14203 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Target criteria: correlation coefficient
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00320227
ELECTRON MICROSCOPYf_angle_d0.77227520
ELECTRON MICROSCOPYf_dihedral_angle_d8.00215985
ELECTRON MICROSCOPYf_chiral_restr0.0463121
ELECTRON MICROSCOPYf_plane_restr0.0063489

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