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- PDB-5lfj: Crystal Structure of the Bacterial Proteasome Activator Bpa of My... -

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Basic information

Entry
Database: PDB / ID: 5lfj
TitleCrystal Structure of the Bacterial Proteasome Activator Bpa of Mycobacterium tuberculosis
ComponentsBacterial proteasome activator
KeywordsCHAPERONE / Dodecamer / four-helix bundle
Function / homologyBacterial proteasome activator / Bacterial proteasome activator / proteasome accessory complex / positive regulation of proteasomal protein catabolic process / proteasome binding / peptidoglycan-based cell wall / protein homooligomerization / plasma membrane / Bacterial proteasome activator
Function and homology information
Biological speciesMycobacterium tuberculosis (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 2.6 Å
AuthorsBolten, M. / Delley, C.L. / Leibundgut, M. / Boehringer, D. / Ban, N. / Weber-Ban, E.
CitationJournal: Structure / Year: 2016
Title: Structural Analysis of the Bacterial Proteasome Activator Bpa in Complex with the 20S Proteasome.
Authors: Marcel Bolten / Cyrille L Delley / Marc Leibundgut / Daniel Boehringer / Nenad Ban / Eilika Weber-Ban /
Abstract: Mycobacterium tuberculosis harbors proteasomes that recruit substrates for degradation through an ubiquitin-like modification pathway. Recently, a non-ATPase activator termed Bpa (bacterial ...Mycobacterium tuberculosis harbors proteasomes that recruit substrates for degradation through an ubiquitin-like modification pathway. Recently, a non-ATPase activator termed Bpa (bacterial proteasome activator) was shown to support an alternate proteasomal degradation pathway. Here, we present the cryo-electron microscopy (cryo-EM) structure of Bpa in complex with the 20S core particle (CP). For docking into the cryo-EM density, we solved the X-ray structure of Bpa, showing that it forms tight four-helix bundles arranged into a 12-membered ring with a 40 Å wide central pore and the C-terminal helix of each protomer protruding from the ring. The Bpa model was fitted into the cryo-EM map of the Bpa-CP complex, revealing its architecture and striking symmetry mismatch. The Bpa-CP interface was resolved to 3.5 Å, showing the interactions between the C-terminal GQYL motif of Bpa and the proteasome α-rings. This docking mode is related to the one observed for eukaryotic activators with features specific to the bacterial complex.
History
DepositionJul 1, 2016Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 23, 2016Provider: repository / Type: Initial release
Revision 1.1Dec 14, 2016Group: Database references
Revision 1.2Jan 10, 2024Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_ncs_dom_lim
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Bacterial proteasome activator
B: Bacterial proteasome activator
C: Bacterial proteasome activator
D: Bacterial proteasome activator


Theoretical massNumber of molelcules
Total (without water)55,5154
Polymers55,5154
Non-polymers00
Water36020
1
A: Bacterial proteasome activator
B: Bacterial proteasome activator
C: Bacterial proteasome activator
D: Bacterial proteasome activator

A: Bacterial proteasome activator
B: Bacterial proteasome activator
C: Bacterial proteasome activator
D: Bacterial proteasome activator

A: Bacterial proteasome activator
B: Bacterial proteasome activator
C: Bacterial proteasome activator
D: Bacterial proteasome activator


Theoretical massNumber of molelcules
Total (without water)166,54612
Polymers166,54612
Non-polymers00
Water21612
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_665-y+1,x-y+1,z1
crystal symmetry operation3_565-x+y,-x+1,z1
Buried area25740 Å2
ΔGint-135 kcal/mol
Surface area62100 Å2
MethodPISA
Unit cell
Length a, b, c (Å)101.378, 101.378, 311.246
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number155
Space group name H-MH32
Components on special symmetry positions
IDModelComponents
11C-206-

HOH

Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11(chain A and (resseq 38:55 or (resid 56 and (name...
21(chain B and (resseq 38:55 or (resid 56 and (name...
31(chain C and (resseq 38:55 or (resid 56 and (name...
41(chain D and (resseq 38:55 or (resid 56 and (name...

NCS domain segments:

Ens-ID: 1

Dom-IDComponent-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDSelection detailsAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
11THRTHRLYSLYS(chain A and (resseq 38:55 or (resid 56 and (name...AA38 - 554 - 21
12GLNGLNGLNGLN(chain A and (resseq 38:55 or (resid 56 and (name...AA5622
13LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
14LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
15LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
16LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
17LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
18LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
19LEULEUARGARG(chain A and (resseq 38:55 or (resid 56 and (name...AA37 - 1453 - 111
21THRTHRLYSLYS(chain B and (resseq 38:55 or (resid 56 and (name...BB38 - 554 - 21
22GLNGLNGLNGLN(chain B and (resseq 38:55 or (resid 56 and (name...BB5622
23SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
24SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
25SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
26SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
27SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
28SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
29SERSERLEULEU(chain B and (resseq 38:55 or (resid 56 and (name...BB36 - 1482 - 114
31THRTHRLYSLYS(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 554 - 21
32GLNGLNGLNGLN(chain C and (resseq 38:55 or (resid 56 and (name...CC5622
33THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
34THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
35THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
36THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
37THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
38THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
39THRTHRALAALA(chain C and (resseq 38:55 or (resid 56 and (name...CC38 - 1444 - 110
41THRTHRLYSLYS(chain D and (resseq 38:55 or (resid 56 and (name...DD38 - 554 - 21
42GLNGLNGLNGLN(chain D and (resseq 38:55 or (resid 56 and (name...DD5622
43LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114
44LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114
45LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114
46LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114
47LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114
48LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114
49LEULEULEULEU(chain D and (resseq 38:55 or (resid 56 and (name...DD37 - 1483 - 114

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Components

#1: Protein
Bacterial proteasome activator


Mass: 13878.801 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria)
Gene: bpa, Rv3780, MTCY13D12.14 / Plasmid: pET24 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Rosetta / References: UniProt: P9WKX3
#2: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 20 / Source method: isolated from a natural source / Formula: H2O

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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: 2.77 Å3/Da / Density % sol: 55.61 %
Crystal growTemperature: 293.15 K / Method: evaporation
Details: 2-4 % (v/v) propane-2-ol, 100 mM HEPES-NaOH pH 7.5-8.2, 150-300 mM MgCl2
PH range: 7.5 - 8.2

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X06SA / Wavelength: 1 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Apr 16, 2015
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 2.6→84.5 Å / Num. obs: 19401 / % possible obs: 100 % / Redundancy: 9.7 % / Biso Wilson estimate: 67.82 Å2 / CC1/2: 0.999 / Rmerge(I) obs: 0.07 / Rsym value: 0.075 / Net I/σ(I): 17.83
Reflection shellResolution: 2.6→2.74 Å / Redundancy: 9.6 % / Rmerge(I) obs: 0.723 / Mean I/σ(I) obs: 2.96 / % possible all: 100

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Phasing

PhasingMethod: molecular replacement

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Processing

Software
NameVersionClassification
XDSOct 15, 2015data reduction
Aimless0.5.23data scaling
PHENIX1.10_2155refinement
PDB_EXTRACT3.2data extraction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5LFP

5lfp


Resolution: 2.6→84.499 Å / SU ML: 0.42 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 34.59
RfactorNum. reflection% reflection
Rfree0.2934 967 5 %
Rwork0.2244 --
obs0.2277 19345 99.62 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso max: 239.14 Å2 / Biso mean: 87.2775 Å2 / Biso min: 35.66 Å2
Refinement stepCycle: final / Resolution: 2.6→84.499 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3467 0 0 20 3487
Biso mean---70.61 -
Num. residues----441
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0083511
X-RAY DIFFRACTIONf_angle_d1.1674751
X-RAY DIFFRACTIONf_chiral_restr0.062554
X-RAY DIFFRACTIONf_plane_restr0.008634
X-RAY DIFFRACTIONf_dihedral_angle_d9.0473032
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDNumberRefine-IDRmsType
11A1991X-RAY DIFFRACTION9.387TORSIONAL
12B1991X-RAY DIFFRACTION9.387TORSIONAL
13C1991X-RAY DIFFRACTION9.387TORSIONAL
14D1991X-RAY DIFFRACTION9.387TORSIONAL
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Total num. of bins used: 7

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
2.6-2.73710.39621350.301225862721100
2.7371-2.90860.38771370.30692574271199
2.9086-3.13320.35911370.276225832720100
3.1332-3.44850.34651360.270726002736100
3.4485-3.94750.31191390.247726282767100
3.9475-4.97340.24441380.180326432781100
4.9734-84.54320.26121450.202327642909100
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
12.6738-0.4973-0.26491.59791.07171.3934-0.2222-1.4707-0.2520.7099-0.3277-0.8164-0.25330.70070.01550.5889-0.0752-0.00720.57010.04850.6711-15.585231.376823.6612
21.1204-0.9887-0.45410.87490.26580.78280.33640.3656-1.03070.5698-0.23880.1809-0.0666-0.7126-0.00280.758-0.0257-0.01320.67670.00890.6084-21.224524.135816.412
31.5309-1.37960.86433.0462-0.71950.4885-0.5609-0.5732-0.34050.80420.33250.46570.6458-0.7863-0.1830.4922-0.05380.23110.45370.2490.9352-34.770828.635126.9227
44.0289-1.18630.88573.8064-2.01251.2770.0858-1.85550.20771.32080.19330.3682-1.27131.11160.04711.0133-0.03560.10151.034-0.00310.6463-23.398937.273931.5408
50.156-0.6338-0.54852.55492.13362.04030.2419-0.7015-0.252-0.3276-0.1518-0.8055-0.4120.13910.2550.2733-0.0203-0.2140.66840.28240.5582-0.020727.166823.8426
61.64160.78180.19171.33460.90810.7816-0.52651.1409-0.9229-0.29760.3664-0.81410.3146-1.40580.00220.69060.0732-0.00320.75590.08670.675-1.13418.137216.4442
70.3127-0.26220.21111.53042.35285.0221-0.1005-1.0108-1.59121.2023-0.51260.53891.1824-0.6118-0.84130.6746-0.07420.04130.49070.35860.9177-15.088715.150926.9772
82.1351-1.62160.51432.90920.4050.5120.3118-1.4808-0.33471.4287-0.1748-0.2663-1.0860.5383-0.26861.1779-0.2401-0.11071.33490.13270.5445-9.905428.391933.782
91.99420.342-0.49251.42190.83791.33960.1172-1.08320.53720.3768-0.0441-0.2624-0.5429-0.3774-0.00530.399-0.0866-0.08840.55620.13040.634615.639931.470823.804
101.0223-0.2270.30221.7070.41730.3612-1.06830.3212-0.663-0.92820.0569-1.49490.9059-0.2911-0.01010.8472-0.1005-0.01610.73870.09710.6819.239423.163216.4404
110.3369-0.08780.17470.59950.29380.30330.2344-0.9209-1.09710.2381-0.29740.69420.9958-0.1731-0.00020.6058-0.0561-0.0730.60520.25810.89898.509413.415426.863
124.61861.20741.68023.8218-0.55591.22270.0639-1.0370.00721.4532-0.07520.1942-1.2849-1.5231-0.00260.97370.0739-0.06060.92020.10750.6576.739127.313230.6355
130.85710.2289-0.89682.06640.67731.404-0.3535-0.26031.03260.8412-0.0327-0.5973-0.21-0.8028-0.00070.4666-0.0147-0.09540.57780.10350.673827.183642.938323.7675
141.0393-0.0439-0.79052.24990.72840.8294-0.66030.13520.1819-0.53560.0276-1.7356-0.2853-0.2716-0.00620.5838-0.1223-0.01390.77350.02850.716634.35737.377816.3904
151.29860.40670.51620.38040.49770.63770.3381-1.6878-1.03830.6246-0.7876-0.98652.0680.3362-0.11990.76730.1014-0.1950.75650.18610.989229.983623.791126.8982
163.2530.89970.63953.1191-0.29650.79140.0507-1.48260.41421.64080.385-0.0608-0.7188-0.53970.0481.122-0.0565-0.07711.1474-0.01570.645421.129835.035833.7243
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1chain 'A' and (resid 44 through 63 )A44 - 63
2X-RAY DIFFRACTION2chain 'A' and (resid 68 through 88 )A68 - 88
3X-RAY DIFFRACTION3chain 'A' and (resid 91 through 100 )A91 - 100
4X-RAY DIFFRACTION4chain 'A' and (resid 112 through 145 )A112 - 145
5X-RAY DIFFRACTION5chain 'B' and (resid 44 through 63 )B44 - 63
6X-RAY DIFFRACTION6chain 'B' and (resid 68 through 88 )B68 - 88
7X-RAY DIFFRACTION7chain 'B' and (resid 91 through 100 )B91 - 100
8X-RAY DIFFRACTION8chain 'B' and (resid 112 through 148 )B112 - 148
9X-RAY DIFFRACTION9chain 'C' and (resid 44 through 63 )C44 - 63
10X-RAY DIFFRACTION10chain 'C' and (resid 68 through 88 )C68 - 88
11X-RAY DIFFRACTION11chain 'C' and (resid 91 through 100 )C91 - 100
12X-RAY DIFFRACTION12chain 'C' and (resid 112 through 144 )C112 - 144
13X-RAY DIFFRACTION13chain 'D' and (resid 44 through 63 )D44 - 63
14X-RAY DIFFRACTION14chain 'D' and (resid 68 through 88 )D68 - 88
15X-RAY DIFFRACTION15chain 'D' and (resid 91 through 100 )D91 - 100
16X-RAY DIFFRACTION16chain 'D' and (resid 112 through 148 )D112 - 148

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