2pfq
From Proteopedia
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==Overview== | ==Overview== | ||
| - | The incorporation of dNMPs into DNA by polymerases involves a phosphoryl | + | The incorporation of dNMPs into DNA by polymerases involves a phosphoryl transfer reaction hypothesized to require two divalent metal ions. Here we investigate this hypothesis using as a model human DNA polymerase lambda (Pol lambda), an enzyme suggested to be activated in vivo by manganese. We report the crystal structures of four complexes of human Pol lambda. In a 1.9 A structure of Pol lambda containing a 3'-OH and the non-hydrolyzable analog dUpnpp, a non-catalytic Na+ ion occupies the site for metal A and the ribose of the primer-terminal nucleotide is found in a conformation that positions the acceptor 3'-OH out of line with the alpha-phosphate and the bridging oxygen of the pyrophosphate leaving group. Soaking this crystal in MnCl2 yielded a 2.0 A structure with Mn2+ occupying the site for metal A. In the presence of Mn2+, the conformation of the ribose is C3'-endo and the 3'-oxygen is in line with the leaving oxygen, at a distance from the phosphorus atom of the alpha-phosphate (3.69 A) consistent with and supporting a catalytic mechanism involving two divalent metal ions. Finally, soaking with MnCl2 converted a pre-catalytic Pol lambda/Na+ complex with unreacted dCTP in the active site into a product complex via catalysis in the crystal. These data provide pre- and post-transition state information and outline in a single crystal the pathway for the phosphoryl transfer reaction carried out by DNA polymerases. |
==About this Structure== | ==About this Structure== | ||
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==Reference== | ==Reference== | ||
| - | Role of the catalytic metal during polymerization by DNA polymerase lambda., Garcia-Diaz M, Bebenek K, Krahn JM, Pedersen LC, Kunkel TA, DNA Repair (Amst). 2007 | + | Role of the catalytic metal during polymerization by DNA polymerase lambda., Garcia-Diaz M, Bebenek K, Krahn JM, Pedersen LC, Kunkel TA, DNA Repair (Amst). 2007 Sep 1;6(9):1333-40. Epub 2007 May 1. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17475573 17475573] |
[[Category: Homo sapiens]] | [[Category: Homo sapiens]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
[[Category: Bebenek, K.]] | [[Category: Bebenek, K.]] | ||
[[Category: Garcia-Diaz, M.]] | [[Category: Garcia-Diaz, M.]] | ||
| - | [[Category: Krahn, J | + | [[Category: Krahn, J M.]] |
| - | [[Category: Kunkel, T | + | [[Category: Kunkel, T A.]] |
| - | [[Category: Pedersen, L | + | [[Category: Pedersen, L C.]] |
[[Category: DCP]] | [[Category: DCP]] | ||
[[Category: MG]] | [[Category: MG]] | ||
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[[Category: phosphoryl transfer reaction]] | [[Category: phosphoryl transfer reaction]] | ||
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 18:28:59 2008'' |
Revision as of 16:28, 21 February 2008
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Manganese promotes catalysis in a DNA polymerase lambda-DNA crystal
Overview
The incorporation of dNMPs into DNA by polymerases involves a phosphoryl transfer reaction hypothesized to require two divalent metal ions. Here we investigate this hypothesis using as a model human DNA polymerase lambda (Pol lambda), an enzyme suggested to be activated in vivo by manganese. We report the crystal structures of four complexes of human Pol lambda. In a 1.9 A structure of Pol lambda containing a 3'-OH and the non-hydrolyzable analog dUpnpp, a non-catalytic Na+ ion occupies the site for metal A and the ribose of the primer-terminal nucleotide is found in a conformation that positions the acceptor 3'-OH out of line with the alpha-phosphate and the bridging oxygen of the pyrophosphate leaving group. Soaking this crystal in MnCl2 yielded a 2.0 A structure with Mn2+ occupying the site for metal A. In the presence of Mn2+, the conformation of the ribose is C3'-endo and the 3'-oxygen is in line with the leaving oxygen, at a distance from the phosphorus atom of the alpha-phosphate (3.69 A) consistent with and supporting a catalytic mechanism involving two divalent metal ions. Finally, soaking with MnCl2 converted a pre-catalytic Pol lambda/Na+ complex with unreacted dCTP in the active site into a product complex via catalysis in the crystal. These data provide pre- and post-transition state information and outline in a single crystal the pathway for the phosphoryl transfer reaction carried out by DNA polymerases.
About this Structure
2PFQ is a Single protein structure of sequence from Homo sapiens with , , , and as ligands. Full crystallographic information is available from OCA.
Reference
Role of the catalytic metal during polymerization by DNA polymerase lambda., Garcia-Diaz M, Bebenek K, Krahn JM, Pedersen LC, Kunkel TA, DNA Repair (Amst). 2007 Sep 1;6(9):1333-40. Epub 2007 May 1. PMID:17475573
Page seeded by OCA on Thu Feb 21 18:28:59 2008
Categories: Homo sapiens | Single protein | Bebenek, K. | Garcia-Diaz, M. | Krahn, J M. | Kunkel, T A. | Pedersen, L C. | DCP | MG | MN | NA | PPV | Dna polymerase | Dna repair | Manganese | Phosphoryl transfer reaction
