Exonuclease III and endonuclease IV remove 3' blocks from DNA synthesis primers in H2O2-damaged Escherichia coli

Abstract

Escherichia coli deficient in exonuclease III (xth gene mutants) are known to be hypersensitive to hydrogen peroxide. We now show that such mutants accumulate many more DNA single-strand breaks than do wild-type bacteria upon exposure to H2O2. DNA isolated from H2O2-treated xth- cells contains strand breaks that do not efficiently support synthesis by E. coli DNA polymerase I, indicating the presence of blocking groups at the DNA 3' termini. Purified E. coli exonuclease III activates this blocked DNA to allow substantial synthesis by polymerase I in vitro. Another E. coli enzyme, endonuclease IV, also activates primers for DNA polymerase. Exonuclease III accounts for greater than 95% of the total activity in E. coli crude extracts for removal of 3'-terminal phosphoglycolaldehyde esters from model DNA substrates. Purified exonuclease III and endonuclease IV can each efficiently remove 3'-terminal phosphoglycolaldehyde in vitro. An important physiological function for exonuclease III is thus the activation of blocked 3' ends for DNA repair synthesis. Endonuclease IV can also initiate the repair of ruptured 3'-deoxyribose in DNA.