Chlamydia pneumoniae AP endonuclease IV could cleave AP sites of double- and single-stranded DNA

Endonuclease IV gene, the only putative AP endonuclease of C. pneumoniae genome, was cloned into pET28a. Recombinant C. pneumoniae endonuclease I V (CpEndoIV) was expressed in E. coli and purified to homogeneity. CpEndoIV has endonuclease activity against apurinic/apyrimidinic sites (AP sites) of double-stranded (ds) oligonucleotides. AP endonuclease activity of CpEndoIV was promoted by divalent metal ions Mg2+ and Zn2+, and inhibited by EDTA. The natural (A, T, C and G) and modified (U, I and 8-oxo-G (GO)) bases opposite AP site had little effect on the cleavage efficiency of AP site of ds oligonucleotides by CpEndoIV. However, the CpEndoIV-dependent cleavage of AP site opposite modified base GO was strongly inhibited by Chlamydia DNA glycosylase MutY. Interestingly, the AP site in single-stranded (ss) oligonucleotides was also the effective substrate of CpEndoIV. Similar to E. coli endonuclease IV, AP endonuclease activity of CpEndoIV was also heat-stable to some extent, with a half time of 5 min at 60 degrees C.