The β Sliding Clamp Binds to Multiple Sites within MutL and MutS

The MutL and MutS proteins are the central components of the DNA repair machinery that corrects mismatches generated by DNA polymerases during synthesis. We find that MutL interacts directly with the β sliding clamp, a ring-shaped dimeric protein that confers processivity to DNA polymerases by tethering them to their substrates. Interestingly, the interaction of MutL with β only occurs in the presence of single-stranded DNA. We find that the interaction occurs via a loop in MutL near the ATP-binding site. The binding site of MutL on β locates to the hydrophobic pocket between domains two and three of the clamp. Site-specific replacement of two residues in MutL diminished interaction with β without disrupting MutL function with helicase II. In vivo studies reveal that this mutant MutL is no longer functional in mismatch repair. In addition, the human MLH1 has a close match to the proliferating cell nuclear antigen clamp binding motif in the region that corresponds to the β interaction site in Escherichia coli MutL, and a peptide corresponding to this site binds proliferating cell nuclear antigen. The current report also examines in detail the interaction of β with MutS. We find that two distinct regions of MutS interact with β. One is located near the C terminus and the other is close to the N terminus, within the mismatch binding domain. Complementation studies using genes encoding different MutS mutants reveal that the N-terminal β interaction motif on MutS is essential for activity in vivo, but the C-terminal interaction site for β is not. In light of these results, we propose roles for the β clamp in orchestrating the sequence of events that lead to mismatch repair in the cell.