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| | ==Structure of bacteriophage T7 lagging-strand DNA polymerase (D5A/E7A) and gp4 (helicase/primase) bound to DNA including RNA/DNA hybrid, and an incoming dTTP (LagS1)== | | ==Structure of bacteriophage T7 lagging-strand DNA polymerase (D5A/E7A) and gp4 (helicase/primase) bound to DNA including RNA/DNA hybrid, and an incoming dTTP (LagS1)== |
| - | <StructureSection load='6n9v' size='340' side='right'caption='[[6n9v]], [[Resolution|resolution]] 4.00Å' scene=''> | + | <SX load='6n9v' size='340' side='right' viewer='molstar' caption='[[6n9v]], [[Resolution|resolution]] 4.00Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[6n9v]] is a 9 chain structure with sequence from [http://en.wikipedia.org/wiki/Bpt7 Bpt7]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6N9V OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6N9V FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[6n9v]] is a 9 chain structure with sequence from [https://en.wikipedia.org/wiki/Escherichia_phage_T7 Escherichia phage T7]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6N9V OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=6N9V FirstGlance]. <br> |
| - | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=TTP:THYMIDINE-5-TRIPHOSPHATE'>TTP</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr> | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">Electron Microscopy, [[Resolution|Resolution]] 4Å</td></tr> |
| - | <tr id='NonStdRes'><td class="sblockLbl"><b>[[Non-Standard_Residue|NonStd Res:]]</b></td><td class="sblockDat"><scene name='pdbligand=DOC:2,3-DIDEOXYCYTIDINE-5-MONOPHOSPHATE'>DOC</scene></td></tr> | + | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=DOC:2,3-DIDEOXYCYTIDINE-5-MONOPHOSPHATE'>DOC</scene>, <scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=TTP:THYMIDINE-5-TRIPHOSPHATE'>TTP</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr> |
| - | <tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[6n7i|6n7i]], [[6n7n|6n7n]], [[6n7s|6n7s]], [[6n7t|6n7t]], [[6n7v|6n7v]], [[6n7w|6n7w]], [[6n9u|6n9u]], [[6n9w|6n9w]], [[6n9x|6n9x]]</td></tr> | + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=6n9v FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6n9v OCA], [https://pdbe.org/6n9v PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=6n9v RCSB], [https://www.ebi.ac.uk/pdbsum/6n9v PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=6n9v ProSAT]</span></td></tr> |
| - | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6n9v FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6n9v OCA], [http://pdbe.org/6n9v PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6n9v RCSB], [http://www.ebi.ac.uk/pdbsum/6n9v PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6n9v ProSAT]</span></td></tr> | + | |
| | </table> | | </table> |
| | == Function == | | == Function == |
| - | [[http://www.uniprot.org/uniprot/PRIM_BPT7 PRIM_BPT7]] Synthesizes short RNA primers for DNA replication. Unwinds the DNA at the replication forks and generates single-stranded DNA for both leading and lagging strand synthesis. The primase synthesizes short RNA primers on the lagging strand that the polymerase elongates using dNTPs.<ref>PMID:9096333</ref> <ref>PMID:21606333</ref> <ref>PMID:22977246</ref> [[http://www.uniprot.org/uniprot/DPOL_BPT7 DPOL_BPT7]] Replicates viral genomic DNA. Non-processive DNA polymerase that achieves processivity by binding to host thioredoxin (TrxA). This interaction increases the rate of dNTP incorporation to yield a processivity of approximately 800 nucleotides (nt) per binding event. Interacts with DNA helicase gp4 to coordinate nucleotide polymerization with unwinding of the DNA. The leading strand is synthesized continuously while synthesis of the lagging strand requires the synthesis of oligoribonucleotides by the primase domain of gp4.<ref>PMID:9218486</ref> <ref>PMID:21606333</ref> | + | [https://www.uniprot.org/uniprot/HELIC_BPT7 HELIC_BPT7] ATP-dependent DNA helicase and primase essential for viral DNA replication and recombination (PubMed:21606333, PubMed:22977246, PubMed:32009150). The helicase moves 5' -> 3' on the lagging strand template, unwinding the DNA duplex ahead of the leading strand polymerase at the replication fork and generating ssDNA for both leading and lagging strand synthesis (PubMed:21606333, PubMed:22977246, PubMed:32009150). ATP or dTTP hydrolysis propels each helicase domain to translocate 2 nt per step sequentially along DNA (PubMed:17604719, PubMed:30679383). Mediates strand transfer when a joint molecule is available and participates in recombinational DNA repair through its role in strand exchange (PubMed:8617248, PubMed:9096333). Primase activity synthesizes short RNA primers at the sequence 5'-GTC-3' on the lagging strand that the polymerase elongates using dNTPs and providing the primase is still present (PubMed:6454135, PubMed:9139692).[HAMAP-Rule:MF_04154]<ref>PMID:17604719</ref> <ref>PMID:21606333</ref> <ref>PMID:22977246</ref> <ref>PMID:30679383</ref> <ref>PMID:32009150</ref> <ref>PMID:6454135</ref> <ref>PMID:8617248</ref> <ref>PMID:9096333</ref> <ref>PMID:9139692</ref> |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
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| | ==See Also== | | ==See Also== |
| | *[[DNA polymerase 3D structures|DNA polymerase 3D structures]] | | *[[DNA polymerase 3D structures|DNA polymerase 3D structures]] |
| | + | *[[RNA polymerase 3D structures|RNA polymerase 3D structures]] |
| | == References == | | == References == |
| | <references/> | | <references/> |
| | __TOC__ | | __TOC__ |
| - | </StructureSection> | + | </SX> |
| - | [[Category: Bpt7]] | + | [[Category: Escherichia phage T7]] |
| | [[Category: Large Structures]] | | [[Category: Large Structures]] |
| - | [[Category: Fox, T]] | + | [[Category: Fox T]] |
| - | [[Category: Gao, Y]] | + | [[Category: Gao Y]] |
| - | [[Category: Val, N]] | + | [[Category: Val N]] |
| - | [[Category: Yang, W]] | + | [[Category: Yang W]] |
| - | [[Category: Dna polymerase]]
| + | |
| - | [[Category: Dna replication]]
| + | |
| - | [[Category: Helicase]]
| + | |
| - | [[Category: Hydrolase]]
| + | |
| - | [[Category: Primase]]
| + | |
| - | [[Category: Replisome]]
| + | |
| - | [[Category: Transferase-dna complex]]
| + | |
| Structural highlights
Function
HELIC_BPT7 ATP-dependent DNA helicase and primase essential for viral DNA replication and recombination (PubMed:21606333, PubMed:22977246, PubMed:32009150). The helicase moves 5' -> 3' on the lagging strand template, unwinding the DNA duplex ahead of the leading strand polymerase at the replication fork and generating ssDNA for both leading and lagging strand synthesis (PubMed:21606333, PubMed:22977246, PubMed:32009150). ATP or dTTP hydrolysis propels each helicase domain to translocate 2 nt per step sequentially along DNA (PubMed:17604719, PubMed:30679383). Mediates strand transfer when a joint molecule is available and participates in recombinational DNA repair through its role in strand exchange (PubMed:8617248, PubMed:9096333). Primase activity synthesizes short RNA primers at the sequence 5'-GTC-3' on the lagging strand that the polymerase elongates using dNTPs and providing the primase is still present (PubMed:6454135, PubMed:9139692).[HAMAP-Rule:MF_04154][1] [2] [3] [4] [5] [6] [7] [8] [9]
Publication Abstract from PubMed
Visualization in atomic detail of the replisome that performs concerted leading- and lagging-DNA strand synthesis at a replication fork has not been reported. Using bacteriophage T7 as a model system, we determined cryo-electron microscopy structures up to 3.2-angstroms resolution of helicase translocating along DNA and of helicase-polymerase-primase complexes engaging in synthesis of both DNA strands. Each domain of the spiral-shaped hexameric helicase translocates sequentially hand-over-hand along a single-stranded DNA coil, akin to the way AAA+ ATPases (adenosine triphosphatases) unfold peptides. Two lagging-strand polymerases are attached to the primase, ready for Okazaki fragment synthesis in tandem. A beta hairpin from the leading-strand polymerase separates two parental DNA strands into a T-shaped fork, thus enabling the closely coupled helicase to advance perpendicular to the downstream DNA duplex. These structures reveal the molecular organization and operating principles of a replisome.
Structures and operating principles of the replisome.,Gao Y, Cui Y, Fox T, Lin S, Wang H, de Val N, Zhou ZH, Yang W Science. 2019 Feb 22;363(6429). pii: science.aav7003. doi:, 10.1126/science.aav7003. Epub 2019 Jan 24. PMID:30679383[10]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Johnson DS, Bai L, Smith BY, Patel SS, Wang MD. Single-molecule studies reveal dynamics of DNA unwinding by the ring-shaped T7 helicase. Cell. 2007 Jun 29;129(7):1299-309. PMID:17604719 doi:10.1016/j.cell.2007.04.038
- ↑ Zhang H, Lee SJ, Zhu B, Tran NQ, Tabor S, Richardson CC. Helicase-DNA polymerase interaction is critical to initiate leading-strand DNA synthesis. Proc Natl Acad Sci U S A. 2011 Jun 7;108(23):9372-7. doi:, 10.1073/pnas.1106678108. Epub 2011 May 23. PMID:21606333 doi:http://dx.doi.org/10.1073/pnas.1106678108
- ↑ Kulczyk AW, Akabayov B, Lee SJ, Bostina M, Berkowitz SA, Richardson CC. An interaction between DNA polymerase and helicase is essential for the high processivity of the bacteriophage T7 replisome. J Biol Chem. 2012 Nov 9;287(46):39050-60. doi: 10.1074/jbc.M112.410647. Epub 2012, Sep 12. PMID:22977246 doi:http://dx.doi.org/10.1074/jbc.M112.410647
- ↑ Gao Y, Cui Y, Fox T, Lin S, Wang H, de Val N, Zhou ZH, Yang W. Structures and operating principles of the replisome. Science. 2019 Feb 22;363(6429). pii: science.aav7003. doi:, 10.1126/science.aav7003. Epub 2019 Jan 24. PMID:30679383 doi:http://dx.doi.org/10.1126/science.aav7003
- ↑ Ma JB, Chen Z, Xu CH, Huang XY, Jia Q, Zou ZY, Mi CY, Ma DF, Lu Y, Zhang HD, Li M. Dynamic structural insights into the molecular mechanism of DNA unwinding by the bacteriophage T7 helicase. Nucleic Acids Res. 2020 Apr 6;48(6):3156-3164. PMID:32009150 doi:10.1093/nar/gkaa057
- ↑ Tabor S, Richardson CC. Template recognition sequence for RNA primer synthesis by gene 4 protein of bacteriophage T7. Proc Natl Acad Sci U S A. 1981 Jan;78(1):205-9. PMID:6454135 doi:10.1073/pnas.78.1.205
- ↑ Kong D, Richardson CC. Single-stranded DNA binding protein and DNA helicase of bacteriophage T7 mediate homologous DNA strand exchange. EMBO J. 1996 Apr 15;15(8):2010-9 PMID:8617248
- ↑ Kong D, Griffith JD, Richardson CC. Gene 4 helicase of bacteriophage T7 mediates strand transfer through pyrimidine dimers, mismatches, and nonhomologous regions. Proc Natl Acad Sci U S A. 1997 Apr 1;94(7):2987-92. PMID:9096333
- ↑ Kusakabe T, Richardson CC. Gene 4 DNA primase of bacteriophage T7 mediates the annealing and extension of ribo-oligonucleotides at primase recognition sites. J Biol Chem. 1997 May 9;272(19):12446-53. PMID:9139692 doi:10.1074/jbc.272.19.12446
- ↑ Gao Y, Cui Y, Fox T, Lin S, Wang H, de Val N, Zhou ZH, Yang W. Structures and operating principles of the replisome. Science. 2019 Feb 22;363(6429). pii: science.aav7003. doi:, 10.1126/science.aav7003. Epub 2019 Jan 24. PMID:30679383 doi:http://dx.doi.org/10.1126/science.aav7003
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