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- PDB-4mq9: Crystal structure of Thermus thermophilus RNA polymerase holoenzy... -

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Basic information

Entry
Database: PDB / ID: 4mq9
TitleCrystal structure of Thermus thermophilus RNA polymerase holoenzyme in complex with GE23077
Components
  • (DNA-directed RNA polymerase subunit ...Polymerase) x 4
  • GE23077
  • RNA polymerase sigma factor
KeywordsTRANSCRIPTION / TRANSFERASE/ANTIBIOTIC / DNA-directed RNA polymerase / transferase-antibiotic complex
Function / homology
Function and homology information


sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding ...sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / metal ion binding / cytoplasm
Similarity search - Function
Four Helix Bundle (Hemerythrin (Met), subunit A) - #1810 / Rna Polymerase Beta Subunit; Chain: C, domain 4 / DNA-directed RNA polymerase, beta subunit, external 1 domain / : / DNA-directed RNA polymerase subunit beta', hybrid domain / Dna-directed Rna Polymerase Ii 140kd Polypeptide; Chain: B; domain 3 / RNA polymerase Rpb2, domain 2 / Dna-directed Rna Polymerase Ii 140kd Polypeptide; Chain: B; Domain 6 / DNA-directed RNA polymerase, subunit 2, domain 6 / RNA polymerase II, Rpb2 subunit, wall domain ...Four Helix Bundle (Hemerythrin (Met), subunit A) - #1810 / Rna Polymerase Beta Subunit; Chain: C, domain 4 / DNA-directed RNA polymerase, beta subunit, external 1 domain / : / DNA-directed RNA polymerase subunit beta', hybrid domain / Dna-directed Rna Polymerase Ii 140kd Polypeptide; Chain: B; domain 3 / RNA polymerase Rpb2, domain 2 / Dna-directed Rna Polymerase Ii 140kd Polypeptide; Chain: B; Domain 6 / DNA-directed RNA polymerase, subunit 2, domain 6 / RNA polymerase II, Rpb2 subunit, wall domain / RNA polymerase subunit, RPB6/omega / Eukaryotic RPB6 RNA polymerase subunit / RNA polymerase II/Efflux pump adaptor protein, barrel-sandwich hybrid domain / DNA-directed RNA polymerase, insert domain / RNA polymerase, RBP11-like subunit / RNA Polymerase Alpha Subunit; Chain A, domain 2 / RNA polymerase sigma factor RpoD, C-terminal / RNA polymerase sigma factor RpoD / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / Sigma-70 factors family signature 2. / RNA polymerase sigma-70 / RNA polymerase sigma-70 region 4 / Sigma-70, region 4 / RNA polymerase sigma-70 region 2 / RNA polymerase sigma-70 like domain / Sigma-70 region 2 / RNA polymerase sigma factor, region 2 / RNA polymerase sigma factor, region 3/4-like / Gyrase A; domain 2 / DNA-directed RNA polymerase, omega subunit / DNA-directed RNA polymerase, subunit beta-prime, bacterial type / DNA-directed RNA polymerase, beta subunit, external 1 domain superfamily / DNA-directed RNA polymerase, beta subunit, external 1 domain / RNA polymerase beta subunit external 1 domain / RNA polymerase, alpha subunit, C-terminal / Bacterial RNA polymerase, alpha chain C terminal domain / DNA-directed RNA polymerase, alpha subunit / DNA-directed RNA polymerase beta subunit, bacterial-type / Four Helix Bundle (Hemerythrin (Met), subunit A) / Beta Complex / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb1, domain 3 superfamily / RNA polymerase Rpb1, clamp domain superfamily / RPB6/omega subunit-like superfamily / DNA-directed RNA polymerase, subunit beta-prime / Winged helix-like DNA-binding domain superfamily/Winged helix DNA-binding domain / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 4 / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 5 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, RBP11-like subunit / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb2, domain 2 / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerases beta chain signature. / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6 / OB fold (Dihydrolipoamide Acetyltransferase, E2P) / Arc Repressor Mutant, subunit A / Roll / Winged helix-like DNA-binding domain superfamily / Alpha-Beta Complex / Up-down Bundle / Beta Barrel / 2-Layer Sandwich / Orthogonal Bundle / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
GE23077 / (2Z)-2-methylbut-2-enoic acid / DNA-directed RNA polymerase subunit alpha / RNA polymerase sigma factor SigA / DNA-directed RNA polymerase subunit omega / DNA-directed RNA polymerase subunit beta' / DNA-directed RNA polymerase subunit beta
Similarity search - Component
Biological speciesThermus thermophilus (bacteria)
Actinomadura sp. (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.35 Å
AuthorsHo, M.X. / Arnold, E. / Ebright, R.H. / Zhang, Y. / Tuske, S.
CitationJournal: Elife / Year: 2014
Title: GE23077 binds to the RNA polymerase 'i' and 'i+1' sites and prevents the binding of initiating nucleotides.
Authors: Zhang, Y. / Degen, D. / Ho, M.X. / Sineva, E. / Ebright, K.Y. / Ebright, Y.W. / Mekler, V. / Vahedian-Movahed, H. / Feng, Y. / Yin, R. / Tuske, S. / Irschik, H. / Jansen, R. / Maffioli, S. / ...Authors: Zhang, Y. / Degen, D. / Ho, M.X. / Sineva, E. / Ebright, K.Y. / Ebright, Y.W. / Mekler, V. / Vahedian-Movahed, H. / Feng, Y. / Yin, R. / Tuske, S. / Irschik, H. / Jansen, R. / Maffioli, S. / Donadio, S. / Arnold, E. / Ebright, R.H.
History
DepositionSep 16, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 7, 2014Provider: repository / Type: Initial release
Revision 1.1May 14, 2014Group: Non-polymer description
Revision 1.2May 28, 2014Group: Derived calculations / Structure summary
Revision 1.3Sep 20, 2023Group: Advisory / Data collection ...Advisory / Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / pdbx_struct_conn_angle / pdbx_unobs_or_zero_occ_atoms / struct_conn / struct_ref_seq_dif
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_comp_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr2_label_atom_id / _pdbx_struct_conn_angle.ptnr2_label_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id / _struct_ref_seq_dif.details

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DNA-directed RNA polymerase subunit alpha
B: DNA-directed RNA polymerase subunit alpha
C: DNA-directed RNA polymerase subunit beta
D: DNA-directed RNA polymerase subunit beta'
E: DNA-directed RNA polymerase subunit omega
F: RNA polymerase sigma factor
I: GE23077
hetero molecules


Theoretical massNumber of molelcules
Total (without water)429,58611
Polymers429,3307
Non-polymers2554
Water724
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area43490 Å2
ΔGint-175 kcal/mol
Surface area136480 Å2
MethodPISA
Unit cell
Length a, b, c (Å)236.569, 236.569, 252.311
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number170
Space group name H-MP65

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Components

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DNA-directed RNA polymerase subunit ... , 4 types, 5 molecules ABCDE

#1: Protein DNA-directed RNA polymerase subunit alpha / Polymerase / RNAP subunit alpha / RNA polymerase subunit alpha / Transcriptase subunit alpha


Mass: 34927.051 Da / Num. of mol.: 2 / Fragment: RPOA / Source method: isolated from a natural source / Source: (natural) Thermus thermophilus (bacteria) / Strain: HB8 / References: UniProt: Q5SHR6, DNA-directed RNA polymerase
#2: Protein DNA-directed RNA polymerase subunit beta / Polymerase / RNAP subunit beta / RNA polymerase subunit beta / Transcriptase subunit beta


Mass: 125436.539 Da / Num. of mol.: 1 / Fragment: RPOB / Source method: isolated from a natural source / Source: (natural) Thermus thermophilus (bacteria) / Strain: HB8 / References: UniProt: Q8RQE9, DNA-directed RNA polymerase
#3: Protein DNA-directed RNA polymerase subunit beta' / Polymerase / RNAP subunit beta' / RNA polymerase subunit beta' / Transcriptase subunit beta'


Mass: 170997.391 Da / Num. of mol.: 1 / Fragment: RPOC / Source method: isolated from a natural source / Source: (natural) Thermus thermophilus (bacteria) / Strain: HB8 / References: UniProt: Q8RQE8, DNA-directed RNA polymerase
#4: Protein DNA-directed RNA polymerase subunit omega / Polymerase / RNAP omega subunit / RNA polymerase omega subunit / Transcriptase subunit omega


Mass: 11533.316 Da / Num. of mol.: 1 / Fragment: RPOZ / Source method: isolated from a natural source / Source: (natural) Thermus thermophilus (bacteria) / Strain: HB8 / References: UniProt: Q8RQE7, DNA-directed RNA polymerase

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Protein / Protein/peptide , 2 types, 2 molecules FI

#5: Protein RNA polymerase sigma factor


Mass: 50769.398 Da / Num. of mol.: 1 / Fragment: RPOD
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermus thermophilus (bacteria) / Strain: HB8 / Gene: RPOD, TTHA0532 / Plasmid: PET28A / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: Q5SKW1, DNA-directed RNA polymerase
#6: Protein/peptide GE23077


Type: Cyclic peptide / Class: Antibiotic, Antimicrobial / Mass: 739.687 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Actinomadura sp. (bacteria) / References: GE23077

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Non-polymers , 4 types, 8 molecules

#7: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
#8: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#9: Chemical ChemComp-MB8 / (2Z)-2-methylbut-2-enoic acid / Angelic acid


Type: Cyclic peptide / Class: Antibiotic, Antimicrobial / Mass: 100.116 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C5H8O2 / References: GE23077
#10: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 4 / 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: 4.75 Å3/Da / Density % sol: 74.09 %
Crystal growTemperature: 295 K / Method: vapor diffusion, hanging drop / pH: 5.4
Details: A stock crystallization solution was prepared by adding T. thermophilus RNAP holoenzyme (10 mg/ml) in 20 mm TRIS-HCl, ph 7.7, 100 mm NaCl, and 1% glycerol to an equal volume of 33 mm ...Details: A stock crystallization solution was prepared by adding T. thermophilus RNAP holoenzyme (10 mg/ml) in 20 mm TRIS-HCl, ph 7.7, 100 mm NaCl, and 1% glycerol to an equal volume of 33 mm magnesium formate containing 40 m ZnCl2. This solution was equilibrated against 35 mm magnesium formate and 30 mm sodium citrate, ph 5.4, vapor diffusion, hanging drop, temperature 295K, VAPOR DIFFUSION, HANGING DROP

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: CHESS / Beamline: F1 / Wavelength: 0.918 / Wavelength: 0.918 Å
DetectorType: ADSC QUANTUM 270 / Detector: CCD / Date: Nov 5, 2010
RadiationMonochromator: HORIZONTAL BENT SI(111), ASYMMETRICALLY CUT WITH WATER COOLED CU BLOCK
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.918 Å / Relative weight: 1
ReflectionResolution: 3.35→39.76 Å / Num. obs: 113506 / % possible obs: 99.1 % / Observed criterion σ(I): 1.35 / Redundancy: 5.81 % / Rmerge(I) obs: 0.112 / Net I/σ(I): 8.5428
Reflection shellResolution: 3.35→3.53 Å / Redundancy: 5.39 % / Rmerge(I) obs: 1 / Mean I/σ(I) obs: 0.76 / % possible all: 97.2

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Processing

Software
NameVersionClassification
PHENIXmodel building
PHENIX(PHENIX.REFINE: 1.7.3_928)refinement
MOSFLMdata reduction
SCALAdata scaling
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 3DXJ

3dxj


Resolution: 3.35→38.971 Å / SU ML: 0.48 / σ(F): 1.35 / Phase error: 29 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.242 2265 2 %
Rwork0.2138 --
obs0.2143 113359 98.9 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 156.78 Å2
Baniso -1Baniso -2Baniso -3
1-16.5497 Å2-0 Å20 Å2
2--16.5497 Å2-0 Å2
3----33.0994 Å2
Refinement stepCycle: LAST / Resolution: 3.35→38.971 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms26543 0 5 4 26552
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00427030
X-RAY DIFFRACTIONf_angle_d0.84636537
X-RAY DIFFRACTIONf_dihedral_angle_d12.77710433
X-RAY DIFFRACTIONf_chiral_restr0.0624123
X-RAY DIFFRACTIONf_plane_restr0.0044798
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.35-3.42280.37841420.3566601X-RAY DIFFRACTION94
3.4228-3.50240.36231560.32576908X-RAY DIFFRACTION99
3.5024-3.58990.33731420.30846958X-RAY DIFFRACTION99
3.5899-3.68690.32391500.28916928X-RAY DIFFRACTION99
3.6869-3.79530.3461090.27567024X-RAY DIFFRACTION100
3.7953-3.91770.31211420.26016952X-RAY DIFFRACTION100
3.9177-4.05760.27721370.24736989X-RAY DIFFRACTION100
4.0576-4.21990.27661220.2297057X-RAY DIFFRACTION100
4.2199-4.41170.25311670.21566908X-RAY DIFFRACTION100
4.4117-4.64390.21771450.20096997X-RAY DIFFRACTION100
4.6439-4.93430.22991390.19647028X-RAY DIFFRACTION100
4.9343-5.31440.25431460.21086990X-RAY DIFFRACTION100
5.3144-5.84760.26031260.21546998X-RAY DIFFRACTION100
5.8476-6.69010.24511410.22296976X-RAY DIFFRACTION99
6.6901-8.41490.21091350.18867000X-RAY DIFFRACTION99
8.4149-38.97350.18251660.1646780X-RAY DIFFRACTION96

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