Cross-utilization of the β sliding clamp by replicative polymerases of evolutionary divergent organisms

Abstract

Chromosomal replicases are multiprotein machines comprised of a DNA polymerase, a sliding clamp, and a clamp loader. This study examines replicase components for their ability to be switched between Gram-positive and Gram-negative organisms. These two cell types diverged over 1 billion years ago, and their sequences have diverged widely. Yet the Escherichia coli β clamp binds directly to Staphylococcus aureus PolC and makes it highly processive, confirming and extending earlier results (Low, R. L., Rashbaum, S. A., and Cozzarelli, N. R. (1976) J. Biol. Chem. 251, 1311-1325). We have also examined the S. aureus β clamp. The results show that it functions with S. aureus PolC, but not with E. coli polymerase III core. PolC is a rather potent polymerase by itself and can extend a primer with an intrinsic speed of 80-120 nucleotides per s. Both E. coli β and S. aureus β converted PolC to a highly processive polymerase, but surprisingly, β also increased the intrinsic rate of DNA synthesis to 240-580 nucleotides per s. This finding expands the scope of β function beyond a simple mechanical tether for processivity to include that of an effector that increases the intrinsic rate of nucleotide incorporation by the polymerase.