Drosophila Rrp1 (Recombination repair protein 1) belongs to a family o
f DNA repair nucleases that includes Escherichia coli exonuclease III,
Streptococcus pneumoniae exonuclease A, bovine BAP, mouse APEX endonu
clease, and human APE. Within a 252 amino acid region, collinear homol
ogy is shared between all members. Rrp1 is unique in that it includes
a 427 amino acid N-terminal region not related to any known sequence.
The protein copurifies with an apurinic endonuclease and a double-stra
nded DNA 3'-exonuclease. In this study, a 5'-end-labeled 37 base pair
oligonucleotide substrate containing a single apurinic site was used t
o characterize the endonuclease activity of Rrp1. This substrate is ut
ilized efficiently by Rrp1: the specific activity observed is 1 x 10(5
) units/mg. The abasic double-stranded DNA oligonucleotide is cleaved
only at the abasic site to create a single-strand break. Strand breaks
are not detected in the complementary strand, in the single-stranded
DNA oligonucleotide, or in the base-paired control substrate. After en
donucleolytic cleavage at the abasic site, exonucleolytic processing a
t the nick is slow and requires a molar excess of Rrp1, while exonucle
ase III degrades the nicked substrate more efficiently. The Rrp1 cleav
age product comigrates with a DNaseI cleavage product, and the newly f
ormed terminus supports DNA synthesis by DNA polymerase. Therefore, Rr
p1 cleaves the phosphodiester backbone at one position 5' to the apuri
nic site and leaves a 3'-hydroxyl terminus. Rrp1 is a class II apurini
c endonuclease and is likely to be important in DNA repair in Drosophi
la.