PDBsum entry 3eql

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protein ligands metals Protein-protein interface(s) links
Transferase PDB id
Protein chains
229 a.a. *
1119 a.a. *
1321 a.a. *
95 a.a. *
345 a.a. *
MXP ×2
_MG ×2
_ZN ×4
Waters ×4496
* Residue conservation analysis
PDB id:
Name: Transferase
Title: Crystal structure of the t. Thermophilus RNA polymerase holoenzyme in complex with antibiotic myxopyronin
Structure: DNA-directed RNA polymerase subunit alpha. Chain: a, b, k, l. Synonym: rnap subunit alpha, transcriptase subunit alpha, RNA polymerase subunit alpha. DNA-directed RNA polymerase subunit beta. Chain: c, m. Synonym: rnap subunit beta, transcriptase subunit beta, RNA polymerase subunit beta. DNA-directed RNA polymerase subunit beta'.
Source: Thermus thermophilus. Organism_taxid: 274. Organism_taxid: 274
2.70Å     R-factor:   0.240     R-free:   0.270
Authors: D.G.Vassylyev,M.N.Vassylyeva,I.Artsimovitch
Key ref:
G.A.Belogurov et al. (2009). Transcription inactivation through local refolding of the RNA polymerase structure. Nature, 457, 332-335. PubMed id: 18946472 DOI: 10.1038/nature07510
30-Sep-08     Release date:   28-Oct-08    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
Q9Z9H6  (RPOA_THETH) -  DNA-directed RNA polymerase subunit alpha
315 a.a.
229 a.a.
Protein chains
Pfam   ArchSchema ?
Q8RQE9  (RPOB_THET8) -  DNA-directed RNA polymerase subunit beta
1119 a.a.
1119 a.a.
Protein chains
Pfam   ArchSchema ?
Q8RQE8  (RPOC_THET8) -  DNA-directed RNA polymerase subunit beta'
1524 a.a.
1321 a.a.
Protein chains
Pfam   ArchSchema ?
Q8RQE7  (RPOZ_THET8) -  DNA-directed RNA polymerase subunit omega
99 a.a.
95 a.a.
Protein chains
Pfam   ArchSchema ?
Q5SKW1  (Q5SKW1_THET8) -  RNA polymerase sigma factor SigA
423 a.a.
345 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: Chains A, B, C, D, E, K, L, M, N, O: E.C.  - DNA-directed Rna polymerase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Nucleoside triphosphate + RNA(n) = diphosphate + RNA(n+1)
Nucleoside triphosphate
+ RNA(n)
= diphosphate
+ RNA(n+1)
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   1 term 
  Biological process     transcription initiation from bacterial-type RNA polymerase promoter   5 terms 
  Biochemical function     transferase activity     9 terms  


DOI no: 10.1038/nature07510 Nature 457:332-335 (2009)
PubMed id: 18946472  
Transcription inactivation through local refolding of the RNA polymerase structure.
G.A.Belogurov, M.N.Vassylyeva, A.Sevostyanova, J.R.Appleman, A.X.Xiang, R.Lira, S.E.Webber, S.Klyuyev, E.Nudler, I.Artsimovitch, D.G.Vassylyev.
Structural studies of antibiotics not only provide a shortcut to medicine allowing for rational structure-based drug design, but may also capture snapshots of dynamic intermediates that become 'frozen' after inhibitor binding. Myxopyronin inhibits bacterial RNA polymerase (RNAP) by an unknown mechanism. Here we report the structure of dMyx-a desmethyl derivative of myxopyronin B-complexed with a Thermus thermophilus RNAP holoenzyme. The antibiotic binds to a pocket deep inside the RNAP clamp head domain, which interacts with the DNA template in the transcription bubble. Notably, binding of dMyx stabilizes refolding of the beta'-subunit switch-2 segment, resulting in a configuration that might indirectly compromise binding to, or directly clash with, the melted template DNA strand. Consistently, footprinting data show that the antibiotic binding does not prevent nucleation of the promoter DNA melting but instead blocks its propagation towards the active site. Myxopyronins are thus, to our knowledge, a first structurally characterized class of antibiotics that target formation of the pre-catalytic transcription initiation complex-the decisive step in gene expression control. Notably, mutations designed in switch-2 mimic the dMyx effects on promoter complexes in the absence of antibiotic. Overall, our results indicate a plausible mechanism of the dMyx action and a stepwise pathway of open complex formation in which core enzyme mediates the final stage of DNA melting near the transcription start site, and that switch-2 might act as a molecular checkpoint for DNA loading in response to regulatory signals or antibiotics. The universally conserved switch-2 may have the same role in all multisubunit RNAPs.
  Selected figure(s)  
Figure 3.
Figure 3: A mechanism of the dMyx action. a, Myx alters the contacts between RNAP and P[R] promoter DNA. A linear DNA fragment encompassing positions -81 to +70 of the P[R] promoter was generated by polymerase chain reaction (PCR); the non-template DNA strand was end-labelled with [^32P]- ATP (see Methods). The sequence from -44 to +23 is shown. The -35 and -10 hexamers are indicated by black boxes, the start site (+1) is shown by a black dot. The top panel shows probing of the non-template strand by piperidine-induced cleavage of the permanganate-modified T residues. Reactivities of -10, -4, -3 and +2 residues (quantification described in Methods) are shown to the left of the gel and summarized above the promoter sequence where black and white arrows indicate high and low reactivity, respectively. The bottom panel shows protection of the non-template DNA strand from DNaseI digestion. The footprint boundaries in the promoter region shown are indicated on the gel and by black (RNAP alone) and white (RNAP with the inhibitor) bars below the promoter sequence; the dideoxy-sequencing ladder is shown for reference. In the gels shown, independent reaction repeats were analysed for consistency. b, Schematic drawing of the putative mechanism of the dMyx action. dwDNA, downstream DNA.
Figure 4.
Figure 4: Mutations in switch-2 affect the open complex formation. Accessibility of the non-template DNA strand residues to permanganate modification probed as in Fig. 3a. Wild-type and mutant RNAPs differ in their patterns of reactivity in the absence of dMyx (top traces) but are nearly identical in the presence of 10 M dMyx (bottom traces). Notably, ' 309–325 that removes the entire rudder loop (which is inserted in the same helix as switch-2, but is unlikely to interfere with the nucleic acids) has no effect on DNA melting, suggesting that a melting defect of a different rudder deletion^19 might be due to changes in the adjacent switch-2 instead.
  The above figures are reprinted from an Open Access publication published by Macmillan Publishers Ltd: Nature (2009, 457, 332-335) copyright 2009.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
  21468230 K.Brodolin (2011).
Antibiotics trapping transcription initiation intermediates: To melt or to bend, what's first?
  Transcription, 2, 60-65.  
  21477256 T.I.Moy, A.Daniel, C.Hardy, A.Jackson, O.Rehrauer, Y.S.Hwang, D.Zou, K.Nguyen, J.A.Silverman, Q.Li, and C.Murphy (2011).
Evaluating the activity of the RNA polymerase inhibitor myxopyronin B against Staphylococcus aureus.
  FEMS Microbiol Lett, 319, 176-179.  
20639538 A.Sevostyanova, and I.Artsimovitch (2010).
Functional analysis of Thermus thermophilus transcription factor NusG.
  Nucleic Acids Res, 38, 7432-7445.  
20562828 A.Tupin, M.Gualtieri, J.P.Leonetti, and K.Brodolin (2010).
The transcription inhibitor lipiarmycin blocks DNA fitting into the RNA polymerase catalytic site.
  EMBO J, 29, 2527-2537.  
20457751 D.Pupov, N.Miropolskaya, A.Sevostyanova, I.Bass, I.Artsimovitch, and A.Kulbachinskiy (2010).
Multiple roles of the RNA polymerase {beta}' SW2 region in transcription initiation, promoter escape, and RNA elongation.
  Nucleic Acids Res, 38, 5784-5796.  
20520915 K.J.Weissman, and R.Müller (2010).
Myxobacterial secondary metabolites: bioactivities and modes-of-action.
  Nat Prod Rep, 27, 1276-1295.  
20503218 O.Erol, T.F.Schäberle, A.Schmitz, S.Rachid, C.Gurgui, M.El Omari, F.Lohr, S.Kehraus, J.Piel, R.Müller, and G.M.König (2010).
Biosynthesis of the myxobacterial antibiotic corallopyronin A.
  Chembiochem, 11, 1253-1265.  
20439713 P.C.Burrows, N.Joly, and M.Buck (2010).
A prehydrolysis state of an AAA+ ATPase supports transcription activation of an enhancer-dependent RNA polymerase.
  Proc Natl Acad Sci U S A, 107, 9376-9381.  
20483995 T.J.Gries, W.S.Kontur, M.W.Capp, R.M.Saecker, and M.T.Record (2010).
One-step DNA melting in the RNA polymerase cleft opens the initiation bubble to form an unstable open complex.
  Proc Natl Acad Sci U S A, 107, 10418-10423.  
19895816 W.J.Lane, and S.A.Darst (2010).
Molecular evolution of multisubunit RNA polymerases: structural analysis.
  J Mol Biol, 395, 686-704.  
19578065 A.Rogozina, E.Zaychikov, M.Buckle, H.Heumann, and B.Sclavi (2009).
DNA melting by RNA polymerase at the T7A1 promoter precedes the rate-limiting step at 37 degrees C and results in the accumulation of an off-pathway intermediate.
  Nucleic Acids Res, 37, 5390-5404.  
19257840 A.Tupin, M.Gualtieri, K.Brodolin, and J.P.Leonetti (2009).
Myxopyronin: a punch in the jaws of bacterial RNA polymerase.
  Future Microbiol, 4, 145-149.  
19489721 J.W.Kozarich (2009).
The biochemistry of disease: desperately seeking syzygy.
  Annu Rev Biochem, 78, 55-63.  
19171784 S.T.Rutherford, C.L.Villers, J.H.Lee, W.Ross, and R.L.Gourse (2009).
Allosteric control of Escherichia coli rRNA promoter complexes by DksA.
  Genes Dev, 23, 236-248.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time.