Abstract
Bacterial chromosome replication is mainly catalyzed by DNA polymerase III, whose beta subunits enable rapid processive DNA replication. Enabled by the clamp-loading complex, the two beta subunits form a ring-like clamp around DNA and keep the polymerase sliding along. Given the essential role of β-clamp, its inhibitors have been explored for antibacterial purposes. Similarly, β-clamp is an ideal target for bacteriophages to shut off host DNA synthesis during host takeover. The Gp168 protein of phage Twort is such an example, which binds to the β-clamp of Staphylococcus aureus and prevents it from loading onto DNA causing replication arrest. Here, we report a cryo-EM structure of the clamp-Gp168 complex at 3.2-Å resolution. In the structure of the complex, the Gp168 dimer occupies the DNA sliding channel of β-clamp and blocks its loading onto DNA, which represents a new inhibitory mechanism against β-clamp function. Interestingly, the key residues responsible for this interaction on the β-clamp are well conserved among bacteria. We therefore demonstrate that Gp168 is potentially a cross-species β-clamp inhibitor, as it forms complex with the Bacillus subtilis β-clamp. Our findings reveal an alternative mechanism for bacteriophages to inhibit β-clamp and provide a new strategy to combat bacterial drug resistance.