DNA-BASE DAMAGE-INDUCED BY IONIZING-RADIATION RECOGNIZED BY ESCHERICHIA-COLI UVRABC NUCLEASE BUT NOT NTH OR FPG PROTEINS

lonizing radiation and other free radical-generating systems induce a great variety of oxidative damage to DNA bases. The major known lesion s are repaired by two well-characterized DNA glycosylases of Escherich ia coli, endonuclease III (Nth) and formamidopyrimidine-DNA glycosylas e (Fpg), which have associated AP lyase activities. To detect and char acterize potentially harmful oxidative base DNA lesions that may be re paired by alternative means, we exposed plasmid DNA to low doses of ga mma-rays and removed the major base lesions by treatment with Nth and Fpg proteins. The closed circular DNA remaining after these treatments was used as a substrate of the UvrABC endonuclease complex from E. co li and as a template in a DNA polymerase arrest assay in vitro. The ci rcular DNA contained lesions that were recognized and incised by the U VrABC nuclease and also lesions that blocked DNA polymerization in vit ro. The blocking lesions were more abundant in DNA irradiated under ni trogen than under air and occurred mainly at tandem guanines; however, they were also frequent at tandem adenines and tandem cytosines. (C) 1996 Wiley-Liss, Inc.