Distinct mechanisms of cis-syn thymine dimer bypass by Dpo4 and DNA polymerase eta

Proc Natl Acad Sci U S A. 2005 Aug 30;102(35):12359-64. doi: 10.1073/pnas.0504380102. Epub 2005 Aug 22.


UV-light-induced cyclobutane pyrimidine dimers (CPDs) present a severe block to synthesis by replicative DNA polymerases (Pols), whereas Poleta promotes proficient and error-free replication through CPDs. Although the archael Dpo4, which, like Poleta, belongs to the Y family of DNA Pols, can also replicate through a CPD, it is much less efficient than Poleta. The x-ray crystal structure of Dpo4 complexed with either the 3'-thymine (T) or the 5' T of a cis-syn TT dimer has indicated that, whereas the 3' T of the dimer forms a Watson-Crick base pair with the incoming dideoxy ATP, the 5' T forms a Hoogsteen base pair with the dideoxy ATP in syn conformation. Based upon these observations, a similar mechanism involving Hoogsteen base pairing of the 5' T of the dimer with the incoming A has been proposed for Poleta. Here we examine the mechanisms of CPD bypass by Dpo4 and Poleta using nucleotide analogs that specifically disrupt the Hoogsteen or Watson-Crick base pairing. Our results show that both Dpo4 and Poleta incorporate dATP opposite the 5' T of the CPD via Watson-Crick base pairing and not by Hoogsteen base pairing. Furthermore, opposite the 3' T of the dimer, the two Pols differ strikingly in the mechanisms of dATP incorporation, with Dpo4 incorporating opposite an abasic-like intermediate and Poleta using the normal Watson-Crick base pairing. These observations have important implications for the mechanisms used for the inefficient vs. efficient bypass of CPDs by DNA Pols.

Publication types

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

MeSH terms

  • Base Pairing
  • Base Sequence
  • Crystallography, X-Ray
  • DNA / chemistry
  • DNA / genetics
  • DNA / metabolism
  • DNA-Directed DNA Polymerase / genetics
  • DNA-Directed DNA Polymerase / metabolism*
  • Humans
  • In Vitro Techniques
  • Kinetics
  • Models, Chemical
  • Molecular Structure
  • Pyrimidine Dimers / chemistry
  • Pyrimidine Dimers / metabolism*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Substrate Specificity
  • Sulfolobus solfataricus / enzymology
  • Sulfolobus solfataricus / genetics


  • Pyrimidine Dimers
  • Recombinant Proteins
  • DNA
  • DNA-Directed DNA Polymerase
  • Rad30 protein