Studies on the catalytic mechanism of five DNA glycosylases. Probing for enzyme-DNA imino intermediates.
DNA glycosylases catalyze scission of the N-glycosylic bond linking a damaged base to the DNA sugar phosphate backbone. Some of these enzymes carry out a concomitant abasic (apyrimidinic/apurinic(AP)) lyase reaction at a rate approximately equal to that of the glycosylase step. As a generalization of the mechanism described for T4 endonuclease V, a repair glycosylase/AP lyase that is specific for ultraviolet light-induced cis-syn pyrimidine dimers, a hypothesis concerning the mechanism of these repair glycosylases has been proposed. This hypothesis describes the initial action of all DNA glycosylases as a nucleophilic attack at the sugar C-1' of the damaged base nucleoside, resulting in scission of the N-glycosylic bond. It is proposed that the enzymes that are only glycosylases differ in the chemical nature of the attacking nucleophile from the glycosylase/AP lyases. Those DNA glycosylases, which carry out the AP lyase reaction at a rate approximately equal to the glycosylase step, are proposed to use an amino group as the nucleophile, resulting in an imino enzyme-DNA intermediate. The simple glycosylases, lacking the concomitant AP lyase activity, are propose to use some nucleophile from the medium, e.g. an activated water molecule. This paper reports experimental tests of this hypothesis using five representative enzymes, and these data are consistent with this hypothesis.
Sun
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Sealy Center for Molecular Science, University of Texas Medical Branch, Galveston 77555-1071, USA.
Latham
K A
KA
Dodson
M L
ML
Lloyd
R S
RS
D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D004247:DNA / Q000378:metabolism*; D045647:DNA Glycosylases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043223:Deoxyribonuclease IV (Phage T4-Induced); D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D007097:Imines / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D008190:Lyases / Q000378:metabolism; D008969:Molecular Sequence Data; D009699:N-Glycosyl Hydrolases / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides
D029968:Escherichia coli Proteins; D007097:Imines; D007527:Isoenzymes; D009838:Oligodeoxyribonucleotides; D004247:DNA; D043223:Deoxyribonuclease IV (Phage T4-Induced); C468972:endonuclease IV, E coli; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D008190:Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase