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PDBsum entry 1msw

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protein dna_rna links
Transcription/DNA/RNA PDB id
1msw
Jmol
Contents
Protein chain
863 a.a. *
DNA/RNA
Waters ×190
* Residue conservation analysis
PDB id:
1msw
Name: Transcription/DNA/RNA
Title: Structural basis for the transition from initiation to elongation transcription in t7 RNA polymerase
Structure: Template DNA. Chain: t. Engineered: yes. Other_details: 20-mer. Non-tempate DNA. Chain: n. Engineered: yes. Other_details: 17-mer. RNA message.
Source: Synthetic: yes. Other_details: this sequence occurs naturally in bacteriophage t7. Enterobacteria phage t7. Organism_taxid: 10760. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.
Biol. unit: Tetramer (from PQS)
Resolution:
2.10Å     R-factor:   0.241     R-free:   0.275
Authors: Y.W.Yin,T.A.Steitz
Key ref:
Y.W.Yin and T.A.Steitz (2002). Structural basis for the transition from initiation to elongation transcription in T7 RNA polymerase. Science, 298, 1387-1395. PubMed id: 12242451 DOI: 10.1126/science.1077464
Date:
19-Sep-02     Release date:   15-Nov-02    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P00573  (RPOL_BPT7) -  T7 RNA polymerase
Seq:
Struc:
 
Seq:
Struc:
883 a.a.
863 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: E.C.2.7.7.6  - 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!
  Biological process     transcription, DNA-dependent   1 term 
  Biochemical function     transferase activity     4 terms  

 

 
    reference    
 
 
DOI no: 10.1126/science.1077464 Science 298:1387-1395 (2002)
PubMed id: 12242451  
 
 
Structural basis for the transition from initiation to elongation transcription in T7 RNA polymerase.
Y.W.Yin, T.A.Steitz.
 
  ABSTRACT  
 
To make messenger RNA transcripts, bacteriophage T7 RNA polymerase (T7 RNAP) undergoes a transition from an initiation phase, which only makes short RNA fragments, to a stable elongation phase. We have determined at 2.1 angstrom resolution the crystal structure of a T7 RNAP elongation complex with 30 base pairs of duplex DNA containing a "transcription bubble" interacting with a 17-nucleotide RNA transcript. The transition from an initiation to an elongation complex is accompanied by a major refolding of the amino-terminal 300 residues. This results in loss of the promoter binding site, facilitating promoter clearance, and creates a tunnel that surrounds the RNA transcript after it peels off a seven-base pair heteroduplex. Formation of the exit tunnel explains the enhanced processivity of the elongation complex. Downstream duplex DNA binds to the fingers domain, and its orientation relative to upstream DNA in the initiation complex implies an unwinding that could facilitate formation of the open promoter complex.
 
  Selected figure(s)  
 
Figure 1.
Fig. 1. Substrates in the T7 RNAP elongation complex. (A) The substrate construct co-crystallized with T7 RNAP consisted of 30 nt each of template DNA (blue) and nontemplate DNA (green) that are complementary (except for a central 11 nt) and a 17-nt RNA (red) whose 3' 10 nt are complementary to the template DNA. The nucleotide that templates the nascent NTP is numbered n and nt upstream are given numbers n i and downstream given numbers n + i (i 1). The portions of RNA and DNA that are not visible in the map are outlined by dashed lines. (B) A portion of the composite omit electron density map corresponding to the substrate is contoured at 1.1 . Figs 1 and 4 were made with the program SPOCK (58).
Figure 3.
Fig. 3. Views of the transcription bubble. (A) Global view of the elongation complex with a box outlining the active site region that is enlarged in (B), (C), and (D) with the thumb (yellow green), subdomain H (green), specificity loop (yellow), and helix Y (red). (B) Conformational changes of the thumb and the specificity loop. The thumb domain as observed in the initiation complex (gray) has rotated about 15° in the elongation complex (green) and assists in the separation of the RNA transcript from the template DNA. The position of the specificity loop in the initiation complex (yellow) blocks the exit of RNA and has moved in the elongation complex (brown) to open the exit tunnel and interact with the exiting RNA. The 3 bp of heteroduplex in the initiation complex (gray) superimposes on that of the elongation complex. (C) Interactions of the transcription bubble and heteroduplex in the elongation complex with domain H (green and red) and specificity loop (brown). Proteolytic cuts within the red loop in subdomain H reduce elongation synthesis (19, 20). Thumb helix (yellow) and -helix Y (orange) are analogously involved in strand separation. (D) Side chains from subdomain H (green), the specificity loop (brown), and the thumb that interact with the single-stranded 5' end of the RNA transcript and facilitate its separation from the template.
 
  The above figures are reprinted by permission from the AAAs: Science (2002, 298, 1387-1395) copyright 2002.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
22245970 D.Takeshita, and K.Tomita (2012).
Molecular basis for RNA polymerization by Qβ replicase.
  Nat Struct Mol Biol, 19, 229-237.
PDB codes: 3avt 3avu 3avv 3avw 3avx 3avy
21947009 R.Ringel, M.Sologub, Y.I.Morozov, D.Litonin, P.Cramer, and D.Temiakov (2011).
Structure of human mitochondrial RNA polymerase.
  Nature, 478, 269-273.
PDB code: 3spa
20067253 G.Stengel, M.Urban, B.W.Purse, and R.D.Kuchta (2010).
Incorporation of the fluorescent ribonucleotide analogue tCTP by T7 RNA polymerase.
  Anal Chem, 82, 1082-1089.  
19297468 A.A.Demidenko, and M.L.Nibert (2009).
Probing the transcription mechanisms of reovirus cores with molecules that alter RNA duplex stability.
  J Virol, 83, 5659-5670.  
19605532 A.C.Rhee, B.H.Somerlot, N.Parimi, and J.M.Gott (2009).
Distinct roles for sequences upstream of and downstream from Physarum editing sites.
  RNA, 15, 1753-1765.  
19801994 C.Tan, P.Marguet, and L.You (2009).
Emergent bistability by a growth-modulating positive feedback circuit.
  Nat Chem Biol, 5, 842-848.  
19896365 D.G.Vassylyev (2009).
Elongation by RNA polymerase: a race through roadblocks.
  Curr Opin Struct Biol, 19, 691-700.  
19015264 E.K.Davydova, I.Kaganman, K.M.Kazmierczak, and L.B.Rothman-Denes (2009).
Identification of bacteriophage N4 virion RNA polymerase-nucleic acid interactions in transcription complexes.
  J Biol Chem, 284, 1962-1970.  
20018723 G.Q.Tang, R.Roy, R.P.Bandwar, T.Ha, and S.S.Patel (2009).
Real-time observation of the transition from transcription initiation to elongation of the RNA polymerase.
  Proc Natl Acad Sci U S A, 106, 22175-22180.  
19239893 H.S.Zaher, and R.Green (2009).
Fidelity at the molecular level: lessons from protein synthesis.
  Cell, 136, 746-762.  
19461581 K.Sugiyama, E.Obayashi, A.Kawaguchi, Y.Suzuki, J.R.Tame, K.Nagata, and S.Y.Park (2009).
Structural insight into the essential PB1-PB2 subunit contact of the influenza virus RNA polymerase.
  EMBO J, 28, 1803-1811.
PDB codes: 2ztt 3a1g
19552410 M.Sorokina, H.R.Koh, S.S.Patel, and T.Ha (2009).
Fluorescent lifetime trajectories of a single fluorophore reveal reaction intermediates during transcription initiation.
  J Am Chem Soc, 131, 9630-9631.  
18022639 A.Dimitri, A.K.Goodenough, F.P.Guengerich, S.Broyde, and D.A.Scicchitano (2008).
Transcription processing at 1,N2-ethenoguanine by human RNA polymerase II and bacteriophage T7 RNA polymerase.
  J Mol Biol, 375, 353-366.  
18854351 A.Dimitri, J.A.Burns, S.Broyde, and D.A.Scicchitano (2008).
Transcription elongation past O6-methylguanine by human RNA polymerase II and bacteriophage T7 RNA polymerase.
  Nucleic Acids Res, 36, 6459-6471.  
18166198 D.Nayak, S.Siller, Q.Guo, and R.Sousa (2008).
Mechanism of T7 RNAP pausing and termination at the T7 concatemer junction: a local change in transcription bubble structure drives a large change in transcription complex architecture.
  J Mol Biol, 376, 541-553.  
18538655 G.Q.Tang, R.Roy, T.Ha, and S.S.Patel (2008).
Transcription initiation in a single-subunit RNA polymerase proceeds through DNA scrunching and rotation of the N-terminal subdomains.
  Mol Cell, 30, 567-577.  
18536012 H.J.Woo, Y.Liu, and R.Sousa (2008).
Molecular dynamics studies of the energetics of translocation in model T7 RNA polymerase elongation complexes.
  Proteins, 73, 1021-1036.  
18070878 K.Datta, and P.H.von Hippel (2008).
Direct spectroscopic study of reconstituted transcription complexes reveals that intrinsic termination is driven primarily by thermodynamic destabilization of the nucleic acid framework.
  J Biol Chem, 283, 3537-3549.  
18948533 K.J.Durniak, S.Bailey, and T.A.Steitz (2008).
The structure of a transcribing T7 RNA polymerase in transition from initiation to elongation.
  Science, 322, 553-557.
PDB codes: 3e2e 3e3j
18362338 K.S.Murakami, E.K.Davydova, and L.B.Rothman-Denes (2008).
X-ray crystal structure of the polymerase domain of the bacteriophage N4 virion RNA polymerase.
  Proc Natl Acad Sci U S A, 105, 5046-5051.
PDB code: 2po4
18834537 L.M.Iyer, S.Abhiman, and L.Aravind (2008).
A new family of polymerases related to superfamily A DNA polymerases and T7-like DNA-dependent RNA polymerases.
  Biol Direct, 3, 39.  
18086892 L.Zhang, A.G.Fletcher, V.Cheung, F.Winston, and L.A.Stargell (2008).
Spn1 regulates the recruitment of Spt6 and the Swi/Snf complex during transcriptional activation by RNA polymerase II.
  Mol Cell Biol, 28, 1393-1403.  
18167548 M.Pandey, S.S.Patel, and A.Gabriel (2008).
Kinetic pathway of pyrophosphorolysis by a retrotransposon reverse transcriptase.
  PLoS ONE, 3, e1389.  
18708471 P.Thomen, P.J.Lopez, U.Bockelmann, J.Guillerez, M.Dreyfus, and F.Heslot (2008).
T7 RNA polymerase studied by force measurements varying cofactor concentration.
  Biophys J, 95, 2423-2433.  
17223130 A.A.Thompson, R.A.Albertini, and O.B.Peersen (2007).
Stabilization of poliovirus polymerase by NTP binding and fingers-thumb interactions.
  J Mol Biol, 366, 1459-1474.
PDB codes: 2ily 2ilz 2im0 2im1 2im2 2im3
17517631 C.Ferrer-Orta, A.Arias, R.Pérez-Luque, C.Escarmís, E.Domingo, and N.Verdaguer (2007).
Sequential structures provide insights into the fidelity of RNA replication.
  Proc Natl Acad Sci U S A, 104, 9463-9468.
PDB codes: 2e9r 2e9t 2e9z 2ec0
17581590 D.G.Vassylyev, M.N.Vassylyeva, A.Perederina, T.H.Tahirov, and I.Artsimovitch (2007).
Structural basis for transcription elongation by bacterial RNA polymerase.
  Nature, 448, 157-162.
PDB code: 2o5i
17580086 D.Nayak, Q.Guo, and R.Sousa (2007).
Functional architecture of T7 RNA polymerase transcription complexes.
  J Mol Biol, 371, 490-500.  
18064834 E.A.Kashkina, M.V.Anikin, W.T.McAllister, N.Kochetkov, and D.E.Temyakov (2007).
Determination of the melting site of the DNA duplex in the active center of bacterial RNA-polymerase by fluorescence quenching technique.
  Dokl Biochem Biophys, 416, 285-289.  
17517766 E.Torreira, G.Schoehn, Y.Fernández, N.Jorba, R.W.Ruigrok, S.Cusack, J.Ortín, and O.Llorca (2007).
Three-dimensional model for the isolated recombinant influenza virus polymerase heterotrimer.
  Nucleic Acids Res, 35, 3774-3783.  
17317624 J.B.Munro, R.B.Altman, N.O'Connor, and S.C.Blanchard (2007).
Identification of two distinct hybrid state intermediates on the ribosome.
  Mol Cell, 25, 505-517.  
17526520 J.H.Kim, and R.G.Larson (2007).
Single-molecule analysis of 1D diffusion and transcription elongation of T7 RNA polymerase along individual stretched DNA molecules.
  Nucleic Acids Res, 35, 3848-3858.  
17400246 P.R.Meyer, W.Rutvisuttinunt, S.E.Matsuura, A.G.So, and W.A.Scott (2007).
Stable complexes formed by HIV-1 reverse transcriptase at distinct positions on the primer-template controlled by binding deoxynucleoside triphosphates or foscarnet.
  J Mol Biol, 369, 41-54.  
17189297 R.K.Shultzaberger, Z.Chen, K.A.Lewis, and T.D.Schneider (2007).
Anatomy of Escherichia coli sigma70 promoters.
  Nucleic Acids Res, 35, 771-788.  
17253774 R.S.Turingan, C.Liu, M.E.Hawkins, and C.T.Martin (2007).
Structural confirmation of a bent and open model for the initiation complex of T7 RNA polymerase.
  Biochemistry, 46, 1714-1723.  
17472344 R.S.Turingan, K.Theis, and C.T.Martin (2007).
Twisted or shifted? Fluorescence measurements of late intermediates in transcription initiation by T7 RNA polymerase.
  Biochemistry, 46, 6165-6168.  
17475199 S.P.Mestas, A.J.Sholders, and O.B.Peersen (2007).
A fluorescence polarization-based screening assay for nucleic acid polymerase elongation activity.
  Anal Biochem, 365, 194-200.  
17553968 Y.Zhou, D.M.Navaroli, M.S.Enuameh, and C.T.Martin (2007).
Dissociation of halted T7 RNA polymerase elongation complexes proceeds via a forward-translocation mechanism.
  Proc Natl Acad Sci U S A, 104, 10352-10357.  
17004274 D.Bednarski, and S.M.Firestine (2006).
Regulation of transcription by synthetic DNA-bending agents.
  Chembiochem, 7, 1715-1721.  
16914440 E.Kashkina, M.Anikin, T.H.Tahirov, S.N.Kochetkov, D.G.Vassylyev, and D.Temiakov (2006).
Elongation complexes of Thermus thermophilus RNA polymerase that possess distinct translocation conformations.
  Nucleic Acids Res, 34, 4036-4045.  
16907127 H.J.Woo (2006).
Analytical theory of the nonequilibrium spatial distribution of RNA polymerase translocations.
  Phys Rev E Stat Nonlin Soft Matter Phys, 74, 011907.  
16765888 J.Zlatanova, W.T.McAllister, S.Borukhov, and S.H.Leuba (2006).
Single-molecule approaches reveal the idiosyncrasies of RNA polymerases.
  Structure, 14, 953-966.  
16565070 K.L.Damm, and H.A.Carlson (2006).
Gaussian-weighted RMSD superposition of proteins: a structural comparison for flexible proteins and predicted protein structures.
  Biophys J, 90, 4558-4573.  
16846227 L.M.Hsu, I.M.Cobb, J.R.Ozmore, M.Khoo, G.Nahm, L.Xia, Y.Bao, and C.Ahn (2006).
Initial transcribed sequence mutations specifically affect promoter escape properties.
  Biochemistry, 45, 8841-8854.  
16674675 N.Crampton, N.H.Thomson, J.Kirkham, C.W.Gibson, and W.A.Bonass (2006).
Imaging RNA polymerase-amelogenin gene complexes with single molecule resolution using atomic force microscopy.
  Eur J Oral Sci, 114, 133.  
17052458 R.T.Pomerantz, D.Temiakov, M.Anikin, D.G.Vassylyev, and W.T.McAllister (2006).
A mechanism of nucleotide misincorporation during transcription due to template-strand misalignment.
  Mol Cell, 24, 245-255.  
16982642 S.D.Auweter, F.C.Oberstrass, and F.H.Allain (2006).
Sequence-specific binding of single-stranded RNA: is there a code for recognition?
  Nucleic Acids Res, 34, 4943-4959.  
16511564 S.Kamtekar, A.J.Berman, J.Wang, J.M.Lázaro, M.de Vega, L.Blanco, M.Salas, and T.A.Steitz (2006).
The phi29 DNA polymerase:protein-primer structure suggests a model for the initiation to elongation transition.
  EMBO J, 25, 1335-1343.
PDB code: 2ex3
16900098 T.A.Steitz (2006).
Visualizing polynucleotide polymerase machines at work.
  EMBO J, 25, 3458-3468.  
15601838 A.Weber, J.Liu, I.Collins, and D.Levens (2005).
TFIIH operates through an expanded proximal promoter to fine-tune c-myc expression.
  Mol Cell Biol, 25, 147-161.  
15574519 F.J.Blocker, G.Mohr, L.H.Conlan, L.Qi, M.Belfort, and A.M.Lambowitz (2005).
Domain structure and three-dimensional model of a group II intron-encoded reverse transcriptase.
  RNA, 11, 14-28.  
15831591 J.Guillerez, P.J.Lopez, F.Proux, H.Launay, and M.Dreyfus (2005).
A mutation in T7 RNA polymerase that facilitates promoter clearance.
  Proc Natl Acad Sci U S A, 102, 5958-5963.  
16301518 K.Ma, D.Temiakov, M.Anikin, and W.T.McAllister (2005).
Probing conformational changes in T7 RNA polymerase during initiation and termination by using engineered disulfide linkages.
  Proc Natl Acad Sci U S A, 102, 17612-17617.  
15903965 P.Thomen, P.J.Lopez, and F.Heslot (2005).
Unravelling the mechanism of RNA-polymerase forward motion by using mechanical force.
  Phys Rev Lett, 94, 128102.  
15722444 V.Alexandrov, U.Lehnert, N.Echols, D.Milburn, D.Engelman, and M.Gerstein (2005).
Normal modes for predicting protein motions: a comprehensive database assessment and associated Web tool.
  Protein Sci, 14, 633-643.  
15914669 Y.Kato, N.Minakawa, Y.Komatsu, H.Kamiya, N.Ogawa, H.Harashima, and A.Matsuda (2005).
New NTP analogs: the synthesis of 4'-thioUTP and 4'-thioCTP and their utility for SELEX.
  Nucleic Acids Res, 33, 2942-2951.  
16094452 Z.F.Burton, M.Feig, X.Q.Gong, C.Zhang, Y.A.Nedialkov, and Y.Xiong (2005).
NTP-driven translocation and regulation of downstream template opening by multi-subunit RNA polymerases.
  Biochem Cell Biol, 83, 486-496.  
15306852 A.A.Thompson, and O.B.Peersen (2004).
Structural basis for proteolysis-dependent activation of the poliovirus RNA-dependent RNA polymerase.
  EMBO J, 23, 3462-3471.
PDB codes: 1ra6 1ra7 1raj 1tql
15016373 D.Temiakov, V.Patlan, M.Anikin, W.T.McAllister, S.Yokoyama, and D.G.Vassylyev (2004).
Structural basis for substrate selection by t7 RNA polymerase.
  Cell, 116, 381-391.
PDB code: 1s0v
15122880 D.W.Gohara, J.J.Arnold, and C.E.Cameron (2004).
Poliovirus RNA-dependent RNA polymerase (3Dpol): kinetic, thermodynamic, and structural analysis of ribonucleotide selection.
  Biochemistry, 43, 5149-5158.  
15610738 H.Kettenberger, K.J.Armache, and P.Cramer (2004).
Complete RNA polymerase II elongation complex structure and its interactions with NTP and TFIIS.
  Mol Cell, 16, 955-965.
PDB codes: 1y1v 1y1w 1y1y 1y77
15109491 I.Artsimovitch, V.Patlan, S.Sekine, M.N.Vassylyeva, T.Hosaka, K.Ochi, S.Yokoyama, and D.G.Vassylyev (2004).
Structural basis for transcription regulation by alarmone ppGpp.
  Cell, 117, 299-310.
PDB codes: 1smy 1t0x
15300257 J.Chelliserrykattil, and A.D.Ellington (2004).
Evolution of a T7 RNA polymerase variant that transcribes 2'-O-methyl RNA.
  Nat Biotechnol, 22, 1155-1160.  
15036151 M.Gerstein, and N.Echols (2004).
Exploring the range of protein flexibility, from a structural proteomics perspective.
  Curr Opin Chem Biol, 8, 14-19.  
15231739 M.L.Duquette, P.Handa, J.A.Vincent, A.F.Taylor, and N.Maizels (2004).
Intracellular transcription of G-rich DNAs induces formation of G-loops, novel structures containing G4 DNA.
  Genes Dev, 18, 1618-1629.  
15196470 P.Cramer (2004).
RNA polymerase II structure: from core to functional complexes.
  Curr Opin Genet Dev, 14, 218-226.  
15035983 S.J.Johnson, and L.S.Beese (2004).
Structures of mismatch replication errors observed in a DNA polymerase.
  Cell, 116, 803-816.
PDB codes: 1njw 1njx 1njy 1njz 1nk0 1nk4 1nk5 1nk6 1nk7 1nk8 1nk9 1nkb 1nkc 1nke
15065652 T.A.Steitz, and Y.W.Yin (2004).
Accuracy, lesion bypass, strand displacement and translocation by DNA polymerases.
  Philos Trans R Soc Lond B Biol Sci, 359, 17-23.  
15102443 T.A.Steitz (2004).
The structural basis of the transition from initiation to elongation phases of transcription, as well as translocation and strand separation, by T7 RNA polymerase.
  Curr Opin Struct Biol, 14, 4-9.  
15016374 Y.W.Yin, and T.A.Steitz (2004).
The structural mechanism of translocation and helicase activity in T7 RNA polymerase.
  Cell, 116, 393-404.
PDB codes: 1s76 1s77
12840000 A.Schwartz, A.R.Rahmouni, and M.Boudvillain (2003).
The functional anatomy of an intrinsic transcription terminator.
  EMBO J, 22, 3385-3394.  
14627736 D.Liu, W.T.Windsor, and D.F.Wyss (2003).
Double-stranded DNA-induced localized unfolding of HCV NS3 helicase subdomain 2.
  Protein Sci, 12, 2757-2767.  
12876194 E.K.Davydova, and L.B.Rothman-Denes (2003).
Escherichia coli single-stranded DNA-binding protein mediates template recycling during transcription by bacteriophage N4 virion RNA polymerase.
  Proc Natl Acad Sci U S A, 100, 9250-9255.  
12581657 K.S.Murakami, and S.A.Darst (2003).
Bacterial RNA polymerases: the wholo story.
  Curr Opin Struct Biol, 13, 31-39.  
12649320 S.J.Johnson, J.S.Taylor, and L.S.Beese (2003).
Processive DNA synthesis observed in a polymerase crystal suggests a mechanism for the prevention of frameshift mutations.
  Proc Natl Acad Sci U S A, 100, 3895-3900.
PDB codes: 1l3s 1l3t 1l3u 1l3v 1l5u 1lv5
14657021 S.Mukherjee, L.G.Brieba, and R.Sousa (2003).
Discontinuous movement and conformational change during pausing and termination by T7 RNA polymerase.
  EMBO J, 22, 6483-6493.  
  12734553 S.Mukherjee, and R.Sousa (2003).
Use of Site-Specifically Tethered Chemical Nucleases to Study Macromolecular Reactions.
  Biol Proced Online, 5, 78-89.  
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. Where a reference describes a PDB structure, the PDB codes are shown on the right.