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| ==Ternary complex of Bacillus DNA Polymerase I Large Fragment, DNA duplex, and rCTP in presence of Mn2+== | | ==Ternary complex of Bacillus DNA Polymerase I Large Fragment, DNA duplex, and rCTP in presence of Mn2+== |
| <StructureSection load='4ds4' size='340' side='right' caption='[[4ds4]], [[Resolution|resolution]] 1.68Å' scene=''> | | <StructureSection load='4ds4' size='340' side='right'caption='[[4ds4]], [[Resolution|resolution]] 1.68Å' scene=''> |
| == Structural highlights == | | == Structural highlights == |
| <table><tr><td colspan='2'>[[4ds4]] is a 6 chain structure with sequence from [http://en.wikipedia.org/wiki/Geoka Geoka]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4DS4 OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4DS4 FirstGlance]. <br> | | <table><tr><td colspan='2'>[[4ds4]] is a 6 chain structure with sequence from [https://en.wikipedia.org/wiki/Geobacillus_kaustophilus_HTA426 Geobacillus kaustophilus HTA426]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4DS4 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4DS4 FirstGlance]. <br> |
| </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=SO4:SULFATE+ION'>SO4</scene></td></tr> | | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.681Å</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=SO4:SULFATE+ION'>SO4</scene></td></tr> |
| <tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4ds5|4ds5]], [[4dse|4dse]], [[4dsf|4dsf]]</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=4ds4 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4ds4 OCA], [https://pdbe.org/4ds4 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4ds4 RCSB], [https://www.ebi.ac.uk/pdbsum/4ds4 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4ds4 ProSAT]</span></td></tr> |
| <tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">polA, GK2730 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=235909 GEOKA])</td></tr>
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| <tr id='activity'><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/DNA-directed_DNA_polymerase DNA-directed DNA polymerase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.7.7.7 2.7.7.7] </span></td></tr>
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| <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=4ds4 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4ds4 OCA], [http://pdbe.org/4ds4 PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=4ds4 RCSB], [http://www.ebi.ac.uk/pdbsum/4ds4 PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=4ds4 ProSAT]</span></td></tr> | |
| </table> | | </table> |
| <div style="background-color:#fffaf0;">
| | == Function == |
| == Publication Abstract from PubMed == | | [https://www.uniprot.org/uniprot/Q5KWC1_GEOKA Q5KWC1_GEOKA] |
| In addition to discriminating against base-pair mismatches, DNA polymerases exhibit a high degree of selectivity for deoxyribonucleotides over ribo- or dideoxy nucleotides. It has been proposed that a single active site residue (steric gate) blocks productive binding of nucleotides containing 2' hydroxyls. Although this steric gate plays a role in sugar moiety discrimination, its interactions do not account fully for the observed behavior of mutants. Here we present ten high-resolution crystal structures and enzyme kinetic analyses of Bacillus DNA polymerase I large fragment (BF) variants complexed with deoxy-, ribo-, dideoxy-nucleotides, and a DNA substrate. Taken together, these data present a more nuanced and general mechanism for nucleotide discrimination in which ensembles of intermediate conformations in the active site trap non-cognate substrates. It is known that the active site O-helix transitions from an open state in the absence of nucleotide substrates to a ternary complex closed state in which the reactive groups are aligned for catalysis. Substrate misalignment in the closed state plays a fundamental part in preventing non-cognate nucleotide misincorpation. The structures presented here show that additional O-helix conformations intermediate between the open and closed state extremes create an ensemble of binding sites that trap and misalign non-cognate nucleotides. Water-mediated interactions, absent in the fully closed state, play an important role in formation of these binding sites, and can be remodeled to accommodate different non-cognate substrates. This mechanism may extend also to base-pair discrimination.
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| Structural factors that determine selectivity of a high-fidelity DNA polymerase for deoxy-, dideoxy-, and ribo-nucleotides.,Wang W, Wu EY, Hellinga HW, Beese LS J Biol Chem. 2012 May 30. PMID:22648417<ref>PMID:22648417</ref>
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| From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br>
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| </div>
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| <div class="pdbe-citations 4ds4" style="background-color:#fffaf0;"></div>
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| ==See Also== | | ==See Also== |
| *[[DNA polymerase|DNA polymerase]] | | *[[DNA polymerase 3D structures|DNA polymerase 3D structures]] |
| == References ==
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| <references/>
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| __TOC__ | | __TOC__ |
| </StructureSection> | | </StructureSection> |
| [[Category: DNA-directed DNA polymerase]] | | [[Category: Geobacillus kaustophilus HTA426]] |
| [[Category: Geoka]] | | [[Category: Large Structures]] |
| [[Category: Beese, L S]] | | [[Category: Beese LS]] |
| [[Category: Wang, W]] | | [[Category: Wang W]] |
| [[Category: Dna polymerase i]]
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| [[Category: Transferase-dna complex]]
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