EXONUCLEASE-I OF ESCHERICHIA-COLI REMOVES PHOSPHOGLYCOLATE 3'-END GROUPS FROM DNA

Exonuclease I of E. coli is a 3' → 5' exonuclease acting on single- stranded DNA. We further demonstrate that the enzyme can remove phosphoglycolate groups at 3' termini in DNA. These types of lesions are introduced into DNA by agents that cause oxidative damage such as ionizing radiation. An oligonucleotide substrate pd(T)20[32P]dA was treated with acid to remove the adenine base to generate 3' termini containing 2- deoxyribose-5-phosphate end groups. This substrate was then treated with periodate to generate 3'-phosphoglycoaldehyde groups and was further oxidized with I2 to generate 3'-phosphoglycolate groups. The pd(T)20[32P]PGA substrate was annealed to pd(A)40-60 to produce a double-stranded substrate. Exonuclease I was effective in the removal of the 3'- phosphoglycolate groups from this substrate as determined by HPLC separation. With exonuclease III and endonuclease IV of E. coli, exonuclease I is the third activity found in E. coli that is able to excise deoxyribose-phosphate fragments at 3' termini in DNA. These sugar fragments are blocks to DNA polymerase, and their removal is necessary to complete the base excision repair process.