1x9s
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T7 DNA polymerase in complex with a primer/template DNA containing a disordered N-2 aminofluorene on the template, crystallized with dideoxy-CTP as the incoming nucleotide.
OverviewOverview
The carcinogen 2-acetylaminofluorene forms two major DNA adducts:, N-(2'-deoxyguanosin-8-yl)-2-acetylaminofluorene (dG-AAF) and its, deacetylated derivative, N-(2'-deoxyguanosin-8-yl)-2-aminofluorene, (dG-AF). Although the dG-AAF and dG-AF adducts are distinguished only by, the presence or absence of an acetyl group, they have profoundly different, effects on DNA replication. dG-AAF poses a strong block to DNA synthesis, and primarily induces frameshift mutations in bacteria, resulting in the, loss of one or two nucleotides during replication past the lesion. dG-AF, is less toxic and more easily bypassed by DNA polymerases, albeit with an, increased frequency of misincorporation opposite the lesion, primarily, resulting in G --> T transversions. We present three crystal structures of, bacteriophage T7 DNA polymerase replication complexes, one with dG-AAF in, the templating position and two others with dG-AF in the templating, position. Our crystallographic data suggest why a dG-AAF adduct blocks, replication more strongly than does a dG-AF adduct and provide a possible, explanation for frameshift mutagenesis during replication bypass of a, dG-AAF adduct. The dG-AAF nucleoside adopts a syn conformation that, facilitates the intercalation of its fluorene ring into a hydrophobic, pocket on the surface of the fingers subdomain and locks the fingers in an, open, inactive conformation. In contrast, the dG-AF base at the templating, position is not well defined by the electron density, consistent with weak, binding to the polymerase and a possible interchange of this adduct, between the syn and anti conformations.
About this StructureAbout this Structure
1X9S is a Protein complex structure of sequences from Bacteriophage t7 and Escherichia coli with MG as ligand. Active as DNA-directed DNA polymerase, with EC number 2.7.7.7 Full crystallographic information is available from OCA.
ReferenceReference
Crystal structures of 2-acetylaminofluorene and 2-aminofluorene in complex with T7 DNA polymerase reveal mechanisms of mutagenesis., Dutta S, Li Y, Johnson D, Dzantiev L, Richardson CC, Romano LJ, Ellenberger T, Proc Natl Acad Sci U S A. 2004 Nov 16;101(46):16186-91. Epub 2004 Nov 4. PMID:15528277
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