Enzymatic release of 5'-terminal deoxyribose phosphate residues from damaged DNA in human cells.
Activities that catalyze or promote the release of 5'-terminal deoxyribose phosphate residues from DNA abasic sites previously incised by an AP endonuclease have been identified in soluble extracts of several human cell lines and calf thymus. Such excision of base-free sugar phosphate residues from apurinic/apyrimidinic sites is expected to be obligatory prior to repair by gap filling and ligation. The most efficient excision function is due to a DNA deoxyribophosphodiesterase similar to the protein found in Escherichia coli. The human enzyme has been partially purified and freed from detectable exonuclease activity. This DNA deoxyribophosphodiesterase is a Mg(2+)-requiring hydrolytic enzyme with an apparent molecular mass of approximately 47 kDa and is located in the cell nucleus. By comparison, the major nuclear 5'----3' exonuclease, DNase IV, is unable to catalyze the release of 5'-terminal deoxyribose phosphate residues as free sugar phosphates but can liberate them at a slow rate as part of small oligonucleotides. Nonenzymatic removal of 5'-terminal deoxyribose phosphate from DNA by beta-elimination promoted by polyamines and basic proteins is a very slow mechanism of release compared to enzymatic hydrolysis. We conclude that a DNA deoxyribophosphodiesterase acts at an intermediate stage between an AP endonuclease and a DNA polymerase during DNA repair at apurinic/apyrimidinc sites in mammalian cells, but several alternative routes also exist for the excision of deoxyribose phosphate residues.
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Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Herts, U.K.
Lindahl
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D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D002417:Cattle; D002460:Cell Line; D004249:DNA Damage; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004706:Endodeoxyribonucleases / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D045585:Flap Endonucleases; D006801:Humans; D007700:Kinetics; D008214:Lymphocytes / Q000201:enzymology*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D012267:Ribosemonophosphates / Q000737:chemistry / Q000378:metabolism*
D029968:Escherichia coli Proteins; D012267:Ribosemonophosphates; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; D045585:Flap Endonucleases; D043223:Deoxyribonuclease IV (Phage T4-Induced); C468972:endonuclease IV, E coli; D010727:Phosphoric Diester Hydrolases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase