Enzymatic release of 5-formyluracil by mammalian liver extracts from DNA irradiated with ionizing radiation.
To identify a repair enzyme for 5-formyluracil (5-FU) caused by ionizing radiation in DNA, we used a radiolabelled product-release assay for this thymine-damaged substrate. Double-stranded poly(dA-dT)-poly(dA-dT) was radiolabelled by nick translation with [2-14C]-thymidine triphosphate. The DNA was irradiated with X-rays and incubated with cell extract from mouse liver. Radiolabelled products released from the irradiated DNA into an ethanol-soluble fraction were analysed by reversed-phase hplc. Released 5-FU was detected as a free base during reaction with the cell extract. 5-Formyl-2'-deoxyuridine was not detected in the ethanol supernatant. Boiling the extract at 97 degrees C for 15 min completely abolished its ability to release 5-FU. Similar enzymatic activity was observed with rat liver extract. These results demonstrated that mammalian cells have enzymatic activity to release 5-FU from DNA.
Zhang
Q M
QM
Laboratory of Radiation Biology, Faculty of Science, Kyoto University, Japan.
Fujimoto
J
J
Yonei
S
S
D000818:Animals; D004247:DNA / Q000378:metabolism / Q000528:radiation effects*; D004260:DNA Repair; D005260:Female; D008099:Liver / Q000378:metabolism*; D051379:Mice; D008813:Mice, Inbred ICR; D010432:Pentoxyl / Q000031:analogs & derivatives / Q000378:metabolism; D051381:Rats; D014020:Tissue Extracts / Q000378:metabolism; D014498:Uracil / Q000031:analogs & derivatives* / Q000378:metabolism
D014020:Tissue Extracts; C007777:5-formyluracil; C008055:5-hydroxymethyluracil; D014498:Uracil; D010432:Pentoxyl; D004247:DNA