Crystal structures of 2-acetylaminofluorene and 2-aminofluorene in complex with T7 DNA polymerase reveal mechanisms of mutagenesis

Proc Natl Acad Sci U S A. 2004 Nov 16;101(46):16186-91. doi: 10.1073/pnas.0406516101. Epub 2004 Nov 4.


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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 2-Acetylaminofluorene / chemistry*
  • 2-Acetylaminofluorene / toxicity
  • Base Sequence
  • Carcinogens / chemistry
  • Carcinogens / toxicity
  • Crystallography, X-Ray
  • DNA / chemistry
  • DNA / genetics
  • DNA Replication
  • DNA-Directed DNA Polymerase / chemistry*
  • Fluorenes / chemistry*
  • Fluorenes / toxicity
  • Macromolecular Substances
  • Models, Molecular
  • Molecular Conformation
  • Mutagenesis


  • Carcinogens
  • Fluorenes
  • Macromolecular Substances
  • 2-aminofluorene
  • DNA
  • 2-Acetylaminofluorene
  • bacteriophage T7 induced DNA polymerase
  • DNA-Directed DNA Polymerase

Associated data

  • PDB/1X9W
  • PDB/IX9M
  • PDB/IX9S