Figure 3.

3′-5′ exonuclease activity of the HhH motif mutants. (A) Exonuclease activity on ssDNA. The assay was performed as described in ‘Materials and Methods’ section, incubating 1.5 nM of the 15mer oligonucleotide with 125 nM of either the wild-type or the indicated mutant PolXBs in the presence of 1 mM of MnCl₂ for the indicated period at 30°C. The unit length of the DNA molecule is indicated. (B) Exonuclease activity on 2-nt gapped DNA. The assay was performed as described in ‘Materials and Methods’ section, incubating 1.5 nM of a 2-nt gapped DNA with 32 nM of either the wild-type or the indicated mutant PolXBs in the presence of 1 mM of MnCl₂ for the indicated period at 30°C. (C) Exonuclease activity on 3′-mismatched gapped substrates. The assay was carried out as described in (B).