Bacteriophage Twort protein Gp168 is a β-clamp inhibitor by occupying the DNA sliding channel

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 beta-clamp, its inhibitors have been explored for antibacterial purposes. Similarly, beta-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 beta-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-angstrom resolution. In the structure of the complex, the Gp168 dimer occupies the DNA sliding channel of beta-clamp and blocks its loading onto DNA, which represents a new inhibitory mechanism against beta-clamp function. Interestingly, the key residues responsible for this interaction on the beta-clamp are well conserved among bacteria. We therefore demonstrate that Gp168 is potentially a cross-species beta-clamp inhibitor, as it forms complex with the Bacillus subtilis beta-clamp. Our findings reveal an alternative mechanism for bacteriophages to inhibit beta-clamp and provide a new strategy to combat bacterial drug resistance.

Automated summary

Bacterial chromosome replication is catalyzed by DNA polymerase III, with the beta subunits of the clamp-loading complex playing a crucial role in enabling processive DNA replication. Inhibitors targeting the beta-clamp have been explored for antibacterial purposes, while bacteriophages like phage Twort can use proteins like Gp168 to bind to the beta-clamp and halt DNA replication. The cryo-EM structure of the clamp-Gp168 complex reveals a new inhibitory mechanism against beta-clamp function, with potential cross-species implications.