Topoisomerases regulate
DNA topology and are fundamental to many aspects of chromosome metabolism. Their activity involves the transient cleavage of
DNA, which, if it occurs near sites of endogenous DNA damage or in the presence of topoisomerase
poisons, can result in abortive topoisomerase-induced
DNA strand breaks. These breaks feature covalent linkage of the
enzyme to the
DNA termini by a 3'- or 5'-phosphotyrosyl bond and are implicated in hereditary human disease,
chromosomal instability and
cancer, and underlie the clinical efficacy of an important class of anti-tumour
poisons. The importance of liberating
DNA termini from trapped topoisomerase is illustrated by the progressive
neurodegenerative disease observed in individuals containing a mutation in
tyrosyl-DNA phosphodiesterase 1 (TDP1), an
enzyme that cleaves 3'-phosphotyrosyl bonds. However, a complementary human
enzyme that cleaves 5'-phosphotyrosyl bonds has not been reported, despite the effect of
DNA double-strand breaks containing such termini on
chromosome instability and
cancer. Here we identify such an
enzyme in human cells and show that this activity efficiently restores 5'-phosphate termini at
DNA double-strand breaks in preparation for
DNA ligation. This
enzyme, TTRAP, is a member of the Mg(2+)/Mn(2+)-dependent family of
phosphodiesterases. Cellular depletion of TTRAP results in increased susceptibility and sensitivity to
topoisomerase-II-induced
DNA double-strand breaks. TTRAP is, to our knowledge, the first human 5'-tyrosyl
DNA phosphodiesterase to be identified, and we suggest that this
enzyme is denoted tyrosyl
DNA phosphodiesterase-2 (TDP2).