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- PDB-6uwi: Crystal structure of the Clostridium difficile translocase CDTb -

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Basic information

Entry
Database: PDB / ID: 6uwi
TitleCrystal structure of the Clostridium difficile translocase CDTb
ComponentsADP-ribosyltransferase binding componentPoly (ADP-ribose) polymerase
KeywordsTOXIN / Translocase
Function / homology
Function and homology information


protein homooligomerization / transferase activity / extracellular region
Similarity search - Function
Bacterial exotoxin B / Protective antigen, heptamerisation domain / Protective antigen, Ca-binding domain / Clostridial binary toxin B/anthrax toxin PA, domain 3 / Protective antigen, heptamerisation domain superfamily / Clostridial binary toxin B/anthrax toxin PA Ca-binding domain / Clostridial binary toxin B/anthrax toxin PA domain 2 / Clostridial binary toxin B/anthrax toxin PA domain 3 / PA14 domain / PA14 / PA14 domain
Similarity search - Domain/homology
ADP-ribosyltransferase binding component
Similarity search - Component
Biological speciesClostridioides difficile (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.7 Å
AuthorsPozharski, E.
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: Structure of the cell-binding component of the binary toxin reveals a di-heptamer macromolecular assembly.
Authors: Xingjian Xu / Raquel Godoy-Ruiz / Kaylin A Adipietro / Christopher Peralta / Danya Ben-Hail / Kristen M Varney / Mary E Cook / Braden M Roth / Paul T Wilder / Thomas Cleveland / Alexander ...Authors: Xingjian Xu / Raquel Godoy-Ruiz / Kaylin A Adipietro / Christopher Peralta / Danya Ben-Hail / Kristen M Varney / Mary E Cook / Braden M Roth / Paul T Wilder / Thomas Cleveland / Alexander Grishaev / Heather M Neu / Sarah L J Michel / Wenbo Yu / Dorothy Beckett / Richard R Rustandi / Catherine Lancaster / John W Loughney / Adam Kristopeit / Sianny Christanti / Jessica W Olson / Alexander D MacKerell / Amedee des Georges / Edwin Pozharski / David J Weber /
Abstract: Targeting infection is challenging because treatment options are limited, and high recurrence rates are common. One reason for this is that hypervirulent strains often have a binary toxin termed ...Targeting infection is challenging because treatment options are limited, and high recurrence rates are common. One reason for this is that hypervirulent strains often have a binary toxin termed the toxin, in addition to the enterotoxins TsdA and TsdB. The toxin has an enzymatic component, termed CDTa, and a pore-forming or delivery subunit termed CDTb. CDTb was characterized here using a combination of single-particle cryoelectron microscopy, X-ray crystallography, NMR, and other biophysical methods. In the absence of CDTa, 2 di-heptamer structures for activated CDTb (1.0 MDa) were solved at atomic resolution, including a symmetric (CDTb; 3.14 Å) and an asymmetric form (CDTb; 2.84 Å). Roles played by 2 receptor-binding domains of activated CDTb were of particular interest since the receptor-binding domain 1 lacks sequence homology to any other known toxin, and the receptor-binding domain 2 is completely absent in other well-studied heptameric toxins (i.e., anthrax). For CDTb, a Ca binding site was discovered in the first receptor-binding domain that is important for its stability, and the second receptor-binding domain was found to be critical for host cell toxicity and the di-heptamer fold for both forms of activated CDTb. Together, these studies represent a starting point for developing structure-based drug-design strategies to target the most severe strains of .
History
DepositionNov 5, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 22, 2020Provider: repository / Type: Initial release
Revision 1.1Mar 6, 2024Group: Data collection / Database references / Derived calculations
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / struct_conn
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: ADP-ribosyltransferase binding component
G: ADP-ribosyltransferase binding component
F: ADP-ribosyltransferase binding component
E: ADP-ribosyltransferase binding component
D: ADP-ribosyltransferase binding component
C: ADP-ribosyltransferase binding component
B: ADP-ribosyltransferase binding component
H: ADP-ribosyltransferase binding component
I: ADP-ribosyltransferase binding component
J: ADP-ribosyltransferase binding component
K: ADP-ribosyltransferase binding component
L: ADP-ribosyltransferase binding component
M: ADP-ribosyltransferase binding component
N: ADP-ribosyltransferase binding component
a: ADP-ribosyltransferase binding component
g: ADP-ribosyltransferase binding component
f: ADP-ribosyltransferase binding component
e: ADP-ribosyltransferase binding component
d: ADP-ribosyltransferase binding component
c: ADP-ribosyltransferase binding component
b: ADP-ribosyltransferase binding component
h: ADP-ribosyltransferase binding component
i: ADP-ribosyltransferase binding component
j: ADP-ribosyltransferase binding component
k: ADP-ribosyltransferase binding component
l: ADP-ribosyltransferase binding component
m: ADP-ribosyltransferase binding component
n: ADP-ribosyltransferase binding component
hetero molecules


Theoretical massNumber of molelcules
Total (without water)2,094,381112
Polymers2,091,01428
Non-polymers3,36784
Water0
1
A: ADP-ribosyltransferase binding component
G: ADP-ribosyltransferase binding component
F: ADP-ribosyltransferase binding component
E: ADP-ribosyltransferase binding component
D: ADP-ribosyltransferase binding component
C: ADP-ribosyltransferase binding component
B: ADP-ribosyltransferase binding component
H: ADP-ribosyltransferase binding component
I: ADP-ribosyltransferase binding component
J: ADP-ribosyltransferase binding component
K: ADP-ribosyltransferase binding component
L: ADP-ribosyltransferase binding component
M: ADP-ribosyltransferase binding component
N: ADP-ribosyltransferase binding component
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,047,19056
Polymers1,045,50714
Non-polymers1,68342
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
a: ADP-ribosyltransferase binding component
g: ADP-ribosyltransferase binding component
f: ADP-ribosyltransferase binding component
e: ADP-ribosyltransferase binding component
d: ADP-ribosyltransferase binding component
c: ADP-ribosyltransferase binding component
b: ADP-ribosyltransferase binding component
h: ADP-ribosyltransferase binding component
i: ADP-ribosyltransferase binding component
j: ADP-ribosyltransferase binding component
k: ADP-ribosyltransferase binding component
l: ADP-ribosyltransferase binding component
m: ADP-ribosyltransferase binding component
n: ADP-ribosyltransferase binding component
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,047,19056
Polymers1,045,50714
Non-polymers1,68342
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)190.485, 190.961, 192.254
Angle α, β, γ (deg.)108.742, 94.473, 108.031
Int Tables number1
Space group name H-MP1
Space group name HallP1
Symmetry operation#1: x,y,z
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-ID
11
21
31
41
51
61
71
81
91
101
111
121
131
141
12
22
32
42
52
62
72
82
92
102
112
122
132
142

NCS domain segments:
Dom-IDComponent-IDEns-IDSelection details
111chain 'A'
211(chain 'B' and (resid 217 through 740 or (resid 741...
311chain 'C'
411(chain 'D' and (resid 217 through 469 or resid 479...
511(chain 'E' and (resid 217 through 469 or resid 479...
611(chain 'F' and (resid 217 through 740 or (resid 741...
711(chain 'G' and (resid 217 through 469 or resid 479...
811chain 'a'
911(chain 'b' and (resid 217 through 740 or (resid 741...
1011chain 'c'
1111(chain 'd' and (resid 217 through 469 or resid 479...
1211(chain 'e' and (resid 217 through 469 or resid 479...
1311(chain 'f' and (resid 217 through 740 or (resid 741...
1411(chain 'g' and (resid 217 through 469 or resid 479...
112chain 'H'
212chain 'I'
312chain 'J'
412chain 'K'
512chain 'L'
612chain 'M'
712chain 'N'
812chain 'h'
912chain 'i'
1012chain 'j'
1112chain 'k'
1212chain 'l'
1312chain 'm'
1412chain 'n'

NCS ensembles :
ID
1
2

NCS oper:
IDCodeMatrixVector
1given(0.786803690539, -0.571433930389, -0.233244969409), (0.331748826822, 0.710222057214, -0.620908483876), (0.520463897362, 0.411154341637, 0.74837787173)21.3312277818, -49.3774913932, -65.668734003
2given(0.302493495833, -0.953149469661, -0.00194254056282), (0.171188645948, 0.056333437666, -0.983626449065), (0.937652458258, 0.297208062283, 0.180208865589)82.6147603193, -36.3617610875, -124.644433784
3given(-0.0824695621373, -0.851229640304, 0.518272969379), (-0.358282238819, -0.459951714834, -0.812452003116), (0.929963767395, -0.252690560743, -0.267048444751)137.180656529, 29.3724375687, -131.358259657
4given(-0.0788584418312, -0.347354539653, 0.934412205579), (-0.856624083685, -0.455807096156, -0.241733469642), (0.509878932103, -0.819502724142, -0.261608026866)143.635945487, 97.9570032386, -81.3706355667
5given(0.310243076037, 0.182158027364, 0.933042167771), (-0.950632372684, 0.066547316299, 0.303099895575), (-0.00687937318663, -0.981014733712, 0.193811161868)97.4629464212, 117.723626857, -12.7669033032
6given(0.788396804198, 0.334503335173, 0.516273181454), (-0.570080131068, 0.712650529929, 0.408824982545), (-0.231169036188, -0.616633392665, 0.752545105464)33.9255621245, 74.3403957205, 23.0318160057
7given(-0.794185125127, 0.560198198418, 0.23547391685), (0.562623226671, 0.531433083404, 0.633272439535), (0.229619450042, 0.635418646527, -0.737236767808)405.295981833, -194.378160755, 110.208584847
8given(-0.999927100624, -0.0112976081544, -0.00426116044957), (-0.00613632518979, 0.171537125155, 0.985158545721), (-0.0103989880061, 0.985112876143, -0.171593946003)425.539868782, -121.206078358, 148.043389025
9given(-0.77843455082, -0.338998540917, -0.528317744682), (-0.331155071, -0.493218482088, 0.804407762194), (-0.533269133778, 0.80113389529, 0.271677221674)390.37656304, -85.8731110763, 217.407709001
10given(-0.298570285523, -0.173865533967, -0.938417050517), (-0.166072552044, -0.958800658239, 0.230480379247), (-0.939827479945, 0.2246599071, 0.257395093352)326.775832461, -115.118151616, 265.185298401
11given(0.0791739951812, 0.35779664763, -0.930437014221), (0.362530007591, -0.879791383062, -0.30747213839), (-0.928602967949, -0.312967540225, -0.199368620104)282.442538843, -186.630937529, 256.036966089
12given(0.0716767354742, 0.860177275003, -0.504933165044), (0.860669270004, -0.309186592915, -0.4045393163), (-0.504094091676, -0.405584400996, -0.762489632983)290.467816796, -247.201520959, 196.463419328
13given(-0.318139657296, 0.947935676799, 0.0143217005421), (0.947980428784, 0.317907324846, 0.0163719104767), (0.0109665445316, 0.0187852458091, -0.99976339673)345.240912226, -250.42775112, 131.661714915
14given(0.78875175672, 0.331765995833, 0.517495884312), (-0.570249459863, 0.70925527911, 0.41445446382), (-0.229534789949, -0.622003434929, 0.748615727286)33.9328366681, 74.2431650374, 22.7760498091
15given(0.301545579557, 0.173090321212, 0.937608662584), (-0.953407727837, 0.045289204848, 0.298265975977), (0.00916340280144, -0.983864131182, 0.178682409381)99.0471351783, 117.239863797, -15.6301020023
16given(-0.0775205556617, -0.356161889104, 0.931203131545), (-0.846206042248, -0.470381065783, -0.25035372379), (0.527186776686, -0.807397276254, -0.264922140986)143.553067747, 95.0508611046, -84.3691602081
17given(-0.0738292501175, -0.850918997076, 0.520082784028), (-0.345843685192, -0.467296887548, -0.813649656978), (0.935383016331, -0.239938490666, -0.259784782959)135.646435993, 26.4945567516, -132.295078191
18given(0.309758170495, -0.950814970716, -0.00087594168323), (0.18244810775, 0.0603423481612, -0.981362058059), (0.933146592873, 0.303825101795, 0.192165927597)81.4525563854, -38.6524172209, -124.276002615
19given(0.789350988038, -0.569152251298, -0.230197160117), (0.334829999385, 0.713366407743, -0.615627517104), (0.514600708506, 0.408869273909, 0.753665726737)20.9375611524, -50.2518483234, -65.2801019869
20given(-0.792003785333, 0.563717526693, 0.234419611205), (0.565763453427, 0.533387844435, 0.628815650391), (0.22943783207, 0.630650424172, -0.741375966502)405.21974215, -194.635176168, 110.114583178
21given(-0.31857176155, 0.947841433954, 0.0104234745996), (0.947895618567, 0.3185232014, 0.00607177662745), (0.00243496296603, 0.0118146624792, -0.999927239706)345.972542587, -249.2232346, 133.16027848
22given(0.074065327531, 0.854354159489, -0.514386330905), (0.84988448581, -0.323930955428, -0.415650209786), (-0.521738141252, -0.406383693423, -0.750094397848)290.817325407, -245.109542532, 199.103845196
23given(0.0743205759329, 0.354976187818, -0.931916497372), (0.347772055792, -0.885059629067, -0.309393035161), (-0.934628829645, -0.301100247555, -0.189228939962)283.914811898, -183.969082388, 257.150158489
24given(-0.302636163108, -0.166663657899, -0.938421322176), (-0.176807800556, -0.957685992735, 0.227104691677), (-0.936563054173, 0.234650302482, 0.260362979518)328.500703521, -112.33199618, 264.846579213
25given(-0.782845995541, -0.333001090675, -0.525606716924), (-0.335965091629, -0.484788652878, 0.807531683123), (-0.523717103476, 0.808758453217, 0.267638113649)391.844044958, -84.4929108239, 215.927263056
26given(-0.999985944604, -0.0043208890893, -0.00307254162561), (-0.00380025094158, 0.180027521185, 0.983654232802), (-0.00369711878956, 0.983652083581, -0.180041411289)426.191117576, -121.106868579, 146.695553327

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Components

#1: Protein ...
ADP-ribosyltransferase binding component / Poly (ADP-ribose) polymerase


Mass: 74679.086 Da / Num. of mol.: 28
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Clostridioides difficile (bacteria) / Gene: cdtB / Production host: Escherichia coli (E. coli) / References: UniProt: O32739
#2: Chemical...
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 84 / Source method: obtained synthetically / Formula: Ca
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.06 Å3/Da / Density % sol: 59.8 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / Details: 22-27% PEG1500, 0.1M MIB, pH 6-8

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRL / Beamline: BL12-2 / Wavelength: 0.97946 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Mar 5, 2017
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97946 Å / Relative weight: 1
ReflectionResolution: 3.7→40 Å / Num. obs: 237933 / % possible obs: 93.2 % / Redundancy: 11.2 % / Biso Wilson estimate: 135.02 Å2 / CC1/2: 0.996 / Net I/σ(I): 4.2
Reflection shellResolution: 3.7→3.76 Å / Redundancy: 9.9 % / Mean I/σ(I) obs: 0.6 / Num. unique obs: 11967 / CC1/2: 0.341 / % possible all: 94.8

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Processing

Software
NameVersionClassification
PHENIX1.17.1_3660refinement
MOSFLMdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: CRYOEM MODEL

Resolution: 3.7→39.75 Å / SU ML: 0.6384 / Cross valid method: FREE R-VALUE / σ(F): 1.96 / Phase error: 32.7033
RfactorNum. reflection% reflection
Rfree0.2727 11423 4.88 %
Rwork0.2485 --
obs0.2497 234142 91.8 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso mean: 127.15 Å2
Refinement stepCycle: LAST / Resolution: 3.7→39.75 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms141578 0 84 0 141662
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0042144112
X-RAY DIFFRACTIONf_angle_d0.7814195598
X-RAY DIFFRACTIONf_chiral_restr0.049422126
X-RAY DIFFRACTIONf_plane_restr0.005225482
X-RAY DIFFRACTIONf_dihedral_angle_d10.99419434
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.7-3.740.40063880.38727298X-RAY DIFFRACTION90.65
3.74-3.790.40673760.37527290X-RAY DIFFRACTION90.49
3.79-3.830.40923820.37717305X-RAY DIFFRACTION89.93
3.83-3.880.41223980.36847230X-RAY DIFFRACTION89.59
3.88-3.930.3653880.35547279X-RAY DIFFRACTION90.09
3.93-3.990.35963790.33947384X-RAY DIFFRACTION91.92
3.99-4.040.38373730.32777376X-RAY DIFFRACTION91.33
4.04-4.10.36243990.32587474X-RAY DIFFRACTION92.38
4.1-4.170.37593540.32687460X-RAY DIFFRACTION91.84
4.17-4.230.34283230.31897434X-RAY DIFFRACTION90.9
4.23-4.310.34023600.31177374X-RAY DIFFRACTION91.36
4.31-4.390.35953930.31487268X-RAY DIFFRACTION89.92
4.39-4.470.34193940.31437146X-RAY DIFFRACTION88.49
4.47-4.560.33333460.30476654X-RAY DIFFRACTION82.08
4.56-4.660.34133640.29767502X-RAY DIFFRACTION92.65
4.66-4.770.29383930.27127765X-RAY DIFFRACTION95.95
4.77-4.890.27783890.25437758X-RAY DIFFRACTION95.93
4.89-5.020.28643830.25447760X-RAY DIFFRACTION95.64
5.02-5.170.26753820.24167703X-RAY DIFFRACTION95.26
5.17-5.330.28324130.25227668X-RAY DIFFRACTION94.86
5.33-5.520.28844290.24227534X-RAY DIFFRACTION94.07
5.52-5.740.27693690.24587656X-RAY DIFFRACTION93.93
5.74-60.27533680.22517510X-RAY DIFFRACTION93.03
6-6.320.24793670.23557342X-RAY DIFFRACTION90.75
6.32-6.710.26863130.24186935X-RAY DIFFRACTION85.2
6.71-7.230.25334100.23077386X-RAY DIFFRACTION91.65
7.23-7.950.22673690.19867780X-RAY DIFFRACTION95.7
7.95-9.090.19294520.17247585X-RAY DIFFRACTION94.7
9.09-11.410.18743750.16357434X-RAY DIFFRACTION91.96
11.41-39.750.20153940.21157429X-RAY DIFFRACTION91.91

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