Selective blockage of the 3 '-> 5 ' exonuclease activity of WRN protein bycertain oxidative modifications and bulky lesions in DNA

Individuals with mutations in the WRN gene suffer from Werner syndrome, a d isease with early onset of many characteristics of normal aging. The WRN pr otein (WRNp) functions in DNA metabolism, as the purified polypeptide has b oth 3'-->5' helicase and 3'-->5' exonuclease activities, In this study, we have further characterized WRNp exonuclease activity by examining its abili ty to degrade double-stranded DNA substrates containing abnormal and damage d nucleotides. In addition, we directly compared the 3'-->5' WRNp exonuclea se activity with that of exonuclease III and the Klenow fragment of DNA pol ymerase I. Our results indicate that the presence of certain abnormal bases (such as uracil and hypoxanthine) does not inhibit the exonuclease activit y of WRNp, exonuclease III or Klenow, whereas other DNA modifications, incl uding apurinic sites, 8-oxoguanine, 8-oxoadenine and cholesterol adducts, i nhibit or block WRNp, The ability of damaged nucleotides to inhibit exonucl eolytic digestion differs significantly between WRNp, exonuclease III and K lenow, indicating that each exonuclease has a distinct mechanism of action, In addition, normal and modified DNA substrates are degraded similarly by full-length WRNp and an N-terminal fragment of WRNp, indicating that the sp ecificity for this activity lies mostly within this region, The biochemical and physiological significance of these results is discussed.