Abstract
Two classes of apurinic/apyrimidinic (AP) endodeoxyribonucleases have been isolated from cultured human fibroblasts. AP endonuclease I cleaves on the 3'-side of the AP site to produce 3'-deoxyribose- and 5'-phosphomonoester termini, which are not efficient primers for Escherichia coli DNA polymerase I. The endonuclease does not activate termini made by preparations of E. coli endonuclease III upon partially depurinated DNA to support DNA synthesis. The latter enzyme also incises the substrate to produce 3'-deoxyribose termini. We have classified AP endonucleases with such a cleavage specificity as Class I. Human fibroblast AP endonuclease II cleaves on the 5'-side of the AP site to produce 3'-hydroxyl nucleotide- and deoxyribose 5-phosphate termini. DNA synthesis occurs from these incisions at a rate similar to that observed from nicks formed by Neurospora crassa endonuclease. We have defined human fibroblast AP endonuclease II as being a member of Class II AP endonucleases, those forming 5'-deoxyribose phosphate termini. AP endonuclease II will activate Class I endonuh AP endonuclease incisions to support DNA synthesis efficiently. The combined actionof AP endonuclease II with AP endonuclease I or with E. coli endonuclease III results in the removal of deoxyribose 5-phosphate from partially depyrimidinated DNA. Whereas normal human fibroblasts contain both APmore » endonucleases, fibroblasts cultured from persons with xeroderma pigmentosum complementation Group D lack AP endonuclease I activity, but contain a normal Class II AP endonuclease. This enzyme deficiency could explain several of the phenotypic characteristics of fibroblasts from this complementation group. As predicted by their specificities, human fibroblasts AP endonuclease II, but not AP endonuclease I can remove the block of E. coli DNA polymerase I DNA synthesis that occurs when T4 uv endonuclease incises DNA substrates containing pyrimidine dimers.« less
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