Partial purification of Pde1 from Saccharomyces cerevisiae: enzymatic redundancy for the repair of 3'-terminal DNA lesions and abasic sites in yeast.
Earlier work indicates that the major DNA repair phosphodiesterase (PDE) in yeast cells is the well-characterized Apn1 protein. Apn1 demonstrates both Mg2+-independent PDE activity and Mg2+-independent class II apurinic/apyrimidinic (AP) endonuclease activity and represents greater than 90% of the activity detected in crude extracts from wild-type yeast cells. Apn1 is related to Echerichia coli endonuclease IV, both in its enzymatic properties and its amino acid sequence. In this work, we report the partial purification of a novel yeast protein, Pde1, present in Apn1-deficient cells. Pde1 is purified by sequential BioRex-70, PBE118, and MonoS chromatography steps using a sensitive and highly specific 3'-phosphoglycolate-terminated oligonucleotide-based assay as a measure of PDE activity. Mg2+-stimulated PDE and Mg2+-stimulated class II AP endonuclease copurify during this procedure. These results indicate that yeast, like many other organisms studied to date, has enzymatic redundancy for the repair of 3'-blocking groups and abasic sites.
Sander
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M
Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709, USA. sander@niehs.nih.gov
Ramotar
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D
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000302:isolation & purification / Q000378:metabolism*; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D004249:DNA Damage; D004260:DNA Repair; D045643:DNA Repair Enzymes; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004706:Endodeoxyribonucleases / Q000172:deficiency; D008190:Lyases / Q000302:isolation & purification / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins
D029701:Saccharomyces cerevisiae Proteins; D004706:Endodeoxyribonucleases; D043223:Deoxyribonuclease IV (Phage T4-Induced); C083274:Apn1 protein, S cerevisiae; D010727:Phosphoric Diester Hydrolases; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D008190:Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D045643:DNA Repair Enzymes