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2imw
From Proteopedia
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(New page: 200px<br /><applet load="2imw" size="450" color="white" frame="true" align="right" spinBox="true" caption="2imw, resolution 2.050Å" /> '''Mechanism of Templa...)
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Revision as of 10:15, 21 November 2007
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Mechanism of Template-Independent Nucleotide Incorporation Catalyzed by a Template-Dependent DNA Polymerase
Overview
Numerous template-dependent DNA polymerases are capable of catalyzing, template-independent nucleotide additions onto blunt-end DNA. Such, non-canonical activity has been hypothesized to increase the genomic, hypermutability of retroviruses including human immunodeficiency viruses., Here, we employed pre-steady state kinetics and X-ray crystallography to, establish a mechanism for blunt-end additions catalyzed by Sulfolobus, solfataricus Dpo4. Our kinetic studies indicated that the first blunt-end, dATP incorporation was 80-fold more efficient than the second, and among, natural deoxynucleotides, dATP was the preferred substrate due to its, stronger intrahelical base-stacking ability. Such base-stacking, contributions are supported by the 41-fold higher ground-state binding, affinity of a nucleotide analog, pyrene nucleoside 5'-triphosphate, which, lacks hydrogen bonding ability but possesses four conjugated aromatic, rings. A 2.05 A resolution structure of Dpo4*(blunt-end DNA)*ddATP, revealed that the base and sugar of the incoming ddATP, respectively, stack against the 5'-base of the opposite strand and the 3'-base of the, elongating strand. This unprecedented base-stacking pattern can be applied, to subsequent blunt-end additions only if all incorporated dAMPs are, extrahelical, leading to predominantly single non-templated dATP, incorporation.
About this Structure
2IMW is a Single protein structure of sequence from Sulfolobus solfataricus with CA, DAD and EDO as ligands. Active as DNA-directed DNA polymerase, with EC number 2.7.7.7 Full crystallographic information is available from OCA.
Reference
Mechanism of template-independent nucleotide incorporation catalyzed by a template-dependent DNA polymerase., Fiala KA, Brown JA, Ling H, Kshetry AK, Zhang J, Taylor JS, Yang W, Suo Z, J Mol Biol. 2007 Jan 19;365(3):590-602. Epub 2006 Oct 7. PMID:17095011
Page seeded by OCA on Wed Nov 21 12:22:50 2007
