Escherichia coli Processivity Clamp β from DNA Polymerase III Is Dynamic in Solution

Escherichia coli DNA polymerase III is a highly processive replicase because of the presence of the beta clamp protein that tethers DNA polymerases to DNA. The beta clamp is a head-to-tail ring-shaped homodimer, in which each protomer contains three structurally similar domains. Although multiple studies have probed the functions of the beta clamp, a detailed understanding of the conformational dynamics of the beta clamp in solution is lacking. Here we used hydrogen exchange mass spectrometry to characterize the conformation and dynamics of the intact dimer beta clamp and a variant form (I272A/L273A) with a weakened ability to dimerize in solution. Our data indicate that the beta clamp is not a static closed ring but rather is dynamic in solution. The three domains exhibited different dynamics, though they share a highly similar tertiary structure. Domain I, which controls the opening of the damp by dissociating from domain III, contained several highly flexible peptides that underwent partial cooperative unfolding (EX1 kinetics) with a half-life of similar to 4 h. The comparison between the beta monomer variant and the wild-type beta clamp showed that the beta monomer was more dynamic. In the monomer, partial unfolding was much faster and additional regions of domain III also underwent partial unfolding with a half-life of similar to 1 h. Our results suggest that the delta subunit of the clamp loader may function as a ring holder to stabilize the transient opening of the beta clamp, rather than as a ring opener.