Purification and properties of the human placental uracil DNA glycosylase.
Human placental uracil DNA glycosylase was purified 3700-fold to apparent homogeneity as defined by SDS gel analysis. Its immunological characteristics were examined using three monoclonal antibodies prepared against partially purified human placental uracil DNA glycosylase. Immunoblot analysis demonstrated that, even in crude isolates, only one glycosylase species of molecular weight 37,000 could be detected. Each of the three monoclonal antibodies quantitatively recognized the highly purified enzyme by ELISA. The glycosylase is a single polypeptide with a molecular weight of 37,000 as defined by both Sephadex gel filtration and by SDS-polyacrylamide gel electrophoresis analysis. The enzyme is heat-stable, with a t 1/2 of greater than 30 min at 42 degrees C or at 45 degrees C. Surprisingly, inhibitor analysis demonstrated that the glycosylase was inhibited by preincubation with either 5-fluorouracil or 5-bromouracil. However, no significant inhibition was observed when either compound was added directly to the enzyme assay.
D000911:Antibodies, Monoclonal; D001976:Bromouracil / Q000494:pharmacology; D002845:Chromatography; D045647:DNA Glycosylases; D004797:Enzyme-Linked Immunosorbent Assay; D005472:Fluorouracil / Q000494:pharmacology; D006358:Hot Temperature; D006801:Humans; D008970:Molecular Weight; D009699:N-Glycosyl Hydrolases / Q000037:antagonists & inhibitors / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D010920:Placenta / Q000201:enzymology*; D051981:Uracil-DNA Glycosidase
D000911:Antibodies, Monoclonal; D001976:Bromouracil; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D051981:Uracil-DNA Glycosidase; D005472:Fluorouracil