Stereoselective excision of thymine glycol from oxidatively damaged DNA.
DNA damage created by reactive oxygen species includes the prototypic oxidized pyrimidine, thymine glycol (Tg), which exists in oxidatively damaged DNA as two diastereoisomeric pairs. In Escherichia coli, Saccharomyces cerevesiae and mice, Tg is preferentially excised by endonuclease III (Endo III) and endonuclease VIII (Endo VIII), yNTG1 and yNTG2, and mNTH and mNEIL1, respectively. We have explored the ability of these DNA glycosylases to discriminate between Tg stereoisomers. Oligonucleotides containing a single, chromatographically pure (5S,6R) or (5R,6S) stereoisomer of Tg were prepared by oxidation with osmium tetroxide. Steady-state kinetic analyses of the excision process revealed that Endo III, Endo VIII, yNTG1, mNTH and mNEIL1, but not yNTG2, excise Tg isomers from DNA in a stereoselective manner, as reflected in the parameter of catalytic efficiency (kcat/Km). When DNA glycosylases occur as complementary pairs, failure of one or both enzymes to excise their cognate Tg stereoisomer from oxidatively damaged DNA could have deleterious consequences for the cell.
Miller
Holly
H
Laboratory of Chemical Biology, State University of New York, Stony Brook, NY, USA. miller@pharm.sunysb.edu
Fernandes
Andrea S
AS
Zaika
Elena
E
McTigue
Monica M
MM
Torres
M Cecilia
MC
Wente
Maryann
M
Iden
Charles R
CR
Grollman
Arthur P
AP
D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004249:DNA Damage; D045647:DNA Glycosylases / Q000378:metabolism*; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043224:Deoxyribonuclease (Pyrimidine Dimer) / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism; D007700:Kinetics; D009699:N-Glycosyl Hydrolases / Q000378:metabolism; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013237:Stereoisomerism; D013379:Substrate Specificity; D013941:Thymine / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000378:metabolism*
D029968:Escherichia coli Proteins; D009841:Oligonucleotides; D029701:Saccharomyces cerevisiae Proteins; C029389:thymine glycol; D004247:DNA; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli; C470703:Nthl1 protein, mouse; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C481347:Neil1 protein, mouse; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; C101825:NTG1 protein, S cerevisiae; C116827:NTG2 protein, S cerevisiae; D013941:Thymine