Mechanism of error-free and semitargeted mutagenic bypass of an aromatic amine lesion by Y-family polymerase Dpo4

Nat Struct Mol Biol. 2010 Mar;17(3):379-88. doi: 10.1038/nsmb.1771. Epub 2010 Feb 14.

Abstract

The aromatic amine carcinogen 2-aminofluorene (AF) forms covalent adducts with DNA, predominantly with guanine at the C8 position. Such lesions are bypassed by Y-family polymerases such as Dpo4 via error-free and error-prone mechanisms. We show that Dpo4 catalyzes elongation from a correct 3'-terminal cytosine opposite [AF]G in a nonrepetitive template sequence with low efficiency. This extension leads to cognate full-length product, as well as mis-elongated products containing base mutations and deletions. Crystal structures of the Dpo4 ternary complex, with the 3'-terminal primer cytosine base opposite [AF]G in the anti conformation and with the AF moiety positioned in the major groove, reveal both accurate and misalignment-mediated mutagenic extension pathways. The mutagenic template-primer-dNTP arrangement is promoted by interactions between the polymerase and the bulky lesion rather than by a base pair-stabilized misaligment. Further extension leads to semitargeted mutations via this proposed polymerase-guided mechanism.

Publication types

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

MeSH terms

  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Crystallography, X-Ray
  • DNA Polymerase beta / chemistry*
  • DNA Polymerase beta / genetics
  • DNA Polymerase beta / metabolism*
  • Fluorenes / chemistry*
  • Fluorenes / metabolism*
  • Kinetics
  • Models, Molecular
  • Protein Structure, Secondary
  • X-Ray Diffraction

Substances

  • Bacterial Proteins
  • Fluorenes
  • 2-aminofluorene
  • DNA Polymerase beta

Associated data

  • PDB/3KHG
  • PDB/3KHH
  • PDB/3KHL
  • PDB/3KHR