Purpose: The aim of this study was to determine the excision efficiency of hSMUG1 (human single-strand-selective monofunctional uracil-DNA glycosylase) for 5-formyluracil (fU), a major thymine lesion formed by ionizing radiation, opposite all normal bases in DNA, to possibly explain mutation induction by fU in the DNA of mammalian cells.
Materials and methods: An enzymatically [(32)P]labelled fU-containing 36 nucleotide DNA sequence plus its complementary sequence (with an A, C, G or T residue inserted opposite fU) was subjected to hSMUG1 in a pH 7.5-buffer, followed by NaOH-mediated cleavage of the resultant abasic sites. Cleaved and uncleaved DNA were separated by denaturing electrophoresis and quantified by autoradiography.
Results: The hSMUG1 excised fU from DNA opposite all normal bases with the highest activity when opposite non-cognate C or T followed by G and cognate A.
Conclusions: The predominant T --> G and T --> A transversions induced by fU in mammalian cells may be explained by replicative incorporation of C and T, respectively, opposite the lesion and subsequent SMUG1-initiated repair of fU.