1x9s: Difference between revisions

Revision as of 16:52, 21 February 2008

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 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

Page seeded by OCA on Thu Feb 21 15:52:29 2008

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-[[Image:1x9s.gif|left|200px]]<br /><applet load="1x9s" size="450" color="white" frame="true" align="right" spinBox="true"
+[[Image:1x9s.gif|left|200px]]<br /><applet load="1x9s" size="350" color="white" frame="true" align="right" spinBox="true"
 caption="1x9s, resolution 2.70&Aring;" />
 '''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.'''<br />
 ==Overview==
-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 --&gt; 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.
+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 --&gt; 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 Structure==
-X9S is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Bacteriophage_t7 Bacteriophage t7] and [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with MG as [http://en.wikipedia.org/wiki/ligand ligand]. 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://ispc.weizmann.ac.il/oca-bin/ocashort?id=1X9S OCA].
+X9S is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Bacteriophage_t7 Bacteriophage t7] and [http://en.wikipedia.org/wiki/Escherichia_coli Escherichia coli] with <scene name='pdbligand=MG:'>MG</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. 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=1X9S OCA].
 ==Reference==
@@ Line 20: / Line 20: @@
 [[Category: Johnson, D.]]
 [[Category: Li, Y.]]
-[[Category: Richardson, C.C.]]
+[[Category: Richardson, C C.]]
-[[Category: Romano, L.J.]]
+[[Category: Romano, L J.]]
 [[Category: MG]]
 [[Category: dna polymerase]]
@@ Line 28: / Line 28: @@
 [[Category: replication block]]
-''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Wed Nov 21 05:55:20 2007''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 15:52:29 2008''