ACTIVITY OF CALF THYMUS DNA HELICASE-E ON CIS-DIAMMINEDICHLOROPLATINUM-(II)-DAMAGED DNA

Calf DNA helicase E (hel E) is a moderately processive, 3' to 5' helic ase, active on nicked DNA, that we have proposed to have a role in DNA repair (Turchi, J. J., Murante, R. S., and Bambara, R. A. (1992) Nucl eic Acids Res. 20, 6075-6080). Here we have examined its activity on a series of cis-diamminedichloroplatinum (II) (cis-DDP)-modified DNA su bstrates. Hel E was capable of efficiently displacing a primer strand containing, in an internal position, a cis-DDP-modified dGG. In a two- primer model system, calf DNA polymerase epsilon could successfully ex tend an upstream primer through a cis-DDP-modified downstream primer, to the end of the complementary template strand, in a reaction depende nt on hel E. However, the translocation of hel E was blocked by cis-DD P modification of the template strand. Primer displacement was complet ely prevented if the modified site was located just upstream of the pr imer. The DNA-dependent ATPase activity of helicase E was also reduced by cis-DDP modification of the template DNA. Substrate competition ex periments indicated that cis-DDP-modified DNA templates did not seques ter hel E. Substrate titration experiments suggested that there is a s hort delay without ATP hydrolysis before dissociation of helicase E fr om cis-DDP-modified template sites. Interestingly, hel E could displac e a primer if the cis-DDP modification was on the template within the annealed region. Possible explanations for this are discussed. Taken t ogether, these results are consistent with the proposal that hel E par ticipates in DNA repair by displacing segments of damaged DNA.