Enzymatic processing of uracil glycol, a major oxidative product of DNA cytosine.
A major stable oxidation product of DNA cytosine is uracil glycol (Ug). Because of the potential of Ug to be a strong premutagenic lesion, it is important to assess whether it is a blocking lesion to DNA polymerase as is its structural counterpart, thymine glycol (Tg), and to evaluate its pairing properties. Here, a series of oligonucleotides containing Ug or Tg were prepared and used as templates for a model enzyme, Escherichia coli DNA polymerase I Klenow fragment (exo-). During translesion DNA synthesis, Ug was bypassed more efficiently than Tg in all sequence contexts examined. Furthermore, only dAMP was incorporated opposite template Ug and Tg and the kinetic parameters of incorporation showed that dAMP was inserted opposite Ug more efficiently than opposite Tg. Ug opposite G and A was also recognized and removed in vitro by the E. coli DNA repair glycosylases, endonuclease III (endo III), endonuclease VIII (endo VIII), and formamidopyrimidine DNA glycosylase. The steady state kinetic parameters indicated that Ug was a better substrate for endo III and formamidopyrimidine DNA glycosylase than Tg; for endonuclease VIII, however, Tg was a better substrate.
Purmal
A A
AA
Department of Microbiology and Molecular Genetics, The Markey Center for Molecular Genetics, University of Vermont, Burlington, Vermont 05405, USA.
Lampman
G W
GW
Bond
J P
JP
Hatahet
Z
Z
Wallace
S S
SS
D000255:Adenosine Triphosphate / Q000378:metabolism; D001482:Base Composition; D001483:Base Sequence; D003596:Cytosine; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004249:DNA Damage; D004256:DNA Polymerase I / Q000378:metabolism*; D017931:DNA Primers; D004260:DNA Repair; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D013698:Templates, Genetic; D013941:Thymine / Q000031:analogs & derivatives / Q000378:metabolism; D014498:Uracil / Q000031:analogs & derivatives* / Q000378:metabolism
D017931:DNA Primers; D003838:Deoxyadenine Nucleotides; D009838:Oligodeoxyribonucleotides; C111900:uracil glycol; C029389:thymine glycol; D014498:Uracil; C012873:2'-deoxy-5'-adenosine monophosphate; D003596:Cytosine; D000255:Adenosine Triphosphate; D004247:DNA; D004256:DNA Polymerase I; D013941:Thymine