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==RNA polymerase II elongation complex with unnatural base dTPT3, rNaM in swing state==
<StructureSection load='7kef' size='340' side='right'caption='[[7kef]], [[Resolution|resolution]] 3.89&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[7kef]] is a 10 chain structure with sequence from [https://en.wikipedia.org/wiki/Saccharomyces_cerevisiae_S288C Saccharomyces cerevisiae S288C]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=7KEF OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=7KEF FirstGlance]. <br>
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 3.89&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=WC4:(1S)-1,4-anhydro-1-(3-methoxynaphthalen-2-yl)-5-O-phosphono-D-ribitol'>WC4</scene>, <scene name='pdbligand=WC7:6-[2-deoxy-5-O-(trihydroxy-lambda~5~-phosphanyl)-beta-D-erythro-pentofuranosyl]thieno[2,3-c]pyridine-7(6H)-thione'>WC7</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></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=7kef FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=7kef OCA], [https://pdbe.org/7kef PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=7kef RCSB], [https://www.ebi.ac.uk/pdbsum/7kef PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=7kef ProSAT]</span></td></tr>
</table>
== Function ==
[https://www.uniprot.org/uniprot/RPB1_YEAST RPB1_YEAST] DNA-dependent RNA polymerase catalyzes the transcription of DNA into RNA using the four ribonucleoside triphosphates as substrates. Largest and catalytic component of RNA polymerase II which synthesizes mRNA precursors and many functional non-coding RNAs. Forms the polymerase active center together with the second largest subunit. Pol II is the central component of the basal RNA polymerase II transcription machinery. During a transcription cycle, Pol II, general transcription factors and the Mediator complex assemble as the preinitiation complex (PIC) at the promoter. 11-15 base pairs of DNA surrounding the transcription start site are melted and the single stranded DNA template strand of the promoter is positioned deeply within the central active site cleft of Pol II to form the open complex. After synthesis of about 30 bases of RNA, Pol II releases its contacts with the core promoter and the rest of the transcription machinery (promoter clearance) and enters the stage of transcription elongation in which it moves on the template as the transcript elongates. Pol II appears to oscillate between inactive and active conformations at each step of nucleotide addition. Elongation is influenced by the phosphorylation status of the C-terminal domain (CTD) of Pol II largest subunit (RPB1), which serves as a platform for assembly of factors that regulate transcription initiation, elongation, termination and mRNA processing. Pol II is composed of mobile elements that move relative to each other. The core element with the central large cleft comprises RPB3, RBP10, RPB11, RPB12 and regions of RPB1 and RPB2 forming the active center. The clamp element (portions of RPB1, RPB2 and RPB3) is connected to the core through a set of flexible switches and moves to open and close the cleft. A bridging helix emanates from RPB1 and crosses the cleft near the catalytic site and is thought to promote translocation of Pol II by acting as a ratchet that moves the RNA-DNA hybrid through the active site by switching from straight to bent conformations at each step of nucleotide addition. In elongating Pol II, the lid loop (RPB1) appears to act as a wedge to drive apart the DNA and RNA strands at the upstream end of the transcription bubble and guide the RNA strand toward the RNA exit groove located near the base of the largely unstructured CTD domain of RPB1. The rudder loop (RPB1) interacts with single stranded DNA after separation from the RNA strand, likely preventing reassociation with the exiting RNA. The cleft is surrounded by jaws: an upper jaw formed by portions of RBP1, RPB2 and RPB9, and a lower jaw, formed by RPB5 and portions of RBP1. The jaws are thought to grab the incoming DNA template, mainly by RPB5 direct contacts to DNA.
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== Publication Abstract from PubMed ==
The development of unnatural base pairs (UBPs) has greatly increased the information storage capacity of DNA, allowing for transcription of unnatural RNA by the heterologously expressed T7 RNA polymerase (RNAP) in Escherichia coli. However, little is known about how UBPs are transcribed by cellular RNA polymerases. Here, we investigated how synthetic unnatural nucleotides, NaM and TPT3, are recognized by eukaryotic RNA polymerase II (Pol II) and found that Pol II is able to selectively recognize UBPs with high fidelity when dTPT3 is in the template strand and rNaMTP acts as the nucleotide substrate. Our structural analysis and molecular dynamics simulation provide structural insights into transcriptional processing of UBPs in a stepwise manner. Intriguingly, we identified a novel 3'-RNA binding site after rNaM addition, termed the swing state. These results may pave the way for future studies in the design of transcription and translation strategies in higher organisms with expanded genetic codes.

Transcriptional processing of an unnatural base pair by eukaryotic RNA polymerase II.,Oh J, Shin J, Unarta IC, Wang W, Feldman AW, Karadeema RJ, Xu L, Xu J, Chong J, Krishnamurthy R, Huang X, Romesberg FE, Wang D Nat Chem Biol. 2021 Aug;17(8):906-914. doi: 10.1038/s41589-021-00817-3. Epub 2021 , Jun 17. PMID:34140682<ref>PMID:34140682</ref>

From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
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==See Also==
*[[RNA polymerase 3D structures|RNA polymerase 3D structures]]
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Large Structures]]
[[Category: Saccharomyces cerevisiae S288C]]
[[Category: Oh J]]
[[Category: Wang D]]