2agq
From Proteopedia
(New page: 200px<br /><applet load="2agq" size="450" color="white" frame="true" align="right" spinBox="true" caption="2agq, resolution 2.10Å" /> '''Fidelity of Dpo4: ef...) |
|||
| Line 1: | Line 1: | ||
| - | [[Image:2agq.gif|left|200px]]<br /><applet load="2agq" size=" | + | [[Image:2agq.gif|left|200px]]<br /><applet load="2agq" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="2agq, resolution 2.10Å" /> | caption="2agq, resolution 2.10Å" /> | ||
'''Fidelity of Dpo4: effect of metal ions, nucleotide selection and pyrophosphorolysis'''<br /> | '''Fidelity of Dpo4: effect of metal ions, nucleotide selection and pyrophosphorolysis'''<br /> | ||
==Overview== | ==Overview== | ||
| - | We report the crystal structures of a translesion DNA polymerase, Dpo4, complexed with a matched or mismatched incoming nucleotide and with a | + | We report the crystal structures of a translesion DNA polymerase, Dpo4, complexed with a matched or mismatched incoming nucleotide and with a pyrophosphate product after misincorporation. These structures suggest two mechanisms by which Dpo4 may reject a wrong incoming nucleotide with its preformed and open active site. First, a mismatched replicating base pair leads to poor base stacking and alignment of the metal ions and as a consequence, inhibits incorporation. By replacing Mg2+ with Mn2+, which has a relaxed coordination requirement and tolerates misalignment, the catalytic efficiency of misincorporation increases dramatically. Mn2+ also enhances translesion synthesis by Dpo4. Subtle conformational changes that lead to the proper metal ion coordination may, therefore, be a key step in catalysis. Second, the slow release of pyrophosphate may increase the fidelity of Dpo4 by stalling mispaired primer extension and promoting pyrophosphorolysis that reverses the polymerization reaction. Indeed, Dpo4 has robust pyrophosphorolysis activity and degrades the primer strand in the presence of pyrophosphate. The correct incoming nucleotide allows DNA synthesis to overcome pyrophosphorolysis, but an incorrect incoming nucleotide does not. |
==About this Structure== | ==About this Structure== | ||
| - | 2AGQ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Sulfolobus_solfataricus Sulfolobus solfataricus] with CA, MG and DTP as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [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] Full crystallographic information is available from [http:// | + | 2AGQ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Sulfolobus_solfataricus Sulfolobus solfataricus] with <scene name='pdbligand=CA:'>CA</scene>, <scene name='pdbligand=MG:'>MG</scene> and <scene name='pdbligand=DTP:'>DTP</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [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] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2AGQ OCA]. |
==Reference== | ==Reference== | ||
| Line 25: | Line 25: | ||
[[Category: pyrophosphorolysis]] | [[Category: pyrophosphorolysis]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:27:14 2008'' |
Revision as of 14:27, 21 February 2008
|
Fidelity of Dpo4: effect of metal ions, nucleotide selection and pyrophosphorolysis
Overview
We report the crystal structures of a translesion DNA polymerase, Dpo4, complexed with a matched or mismatched incoming nucleotide and with a pyrophosphate product after misincorporation. These structures suggest two mechanisms by which Dpo4 may reject a wrong incoming nucleotide with its preformed and open active site. First, a mismatched replicating base pair leads to poor base stacking and alignment of the metal ions and as a consequence, inhibits incorporation. By replacing Mg2+ with Mn2+, which has a relaxed coordination requirement and tolerates misalignment, the catalytic efficiency of misincorporation increases dramatically. Mn2+ also enhances translesion synthesis by Dpo4. Subtle conformational changes that lead to the proper metal ion coordination may, therefore, be a key step in catalysis. Second, the slow release of pyrophosphate may increase the fidelity of Dpo4 by stalling mispaired primer extension and promoting pyrophosphorolysis that reverses the polymerization reaction. Indeed, Dpo4 has robust pyrophosphorolysis activity and degrades the primer strand in the presence of pyrophosphate. The correct incoming nucleotide allows DNA synthesis to overcome pyrophosphorolysis, but an incorrect incoming nucleotide does not.
About this Structure
2AGQ is a Single protein structure of sequence from Sulfolobus solfataricus with , and as ligands. Active as DNA-directed DNA polymerase, with EC number 2.7.7.7 Full crystallographic information is available from OCA.
Reference
Fidelity of Dpo4: effect of metal ions, nucleotide selection and pyrophosphorolysis., Vaisman A, Ling H, Woodgate R, Yang W, EMBO J. 2005 Sep 7;24(17):2957-67. Epub 2005 Aug 18. PMID:16107880
Page seeded by OCA on Thu Feb 21 16:27:14 2008
