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Bacterial proteins are synthesized with an N-formylated amino-terminal methionine, and N-formylated peptides elicit innate-immunity responses against bacterial infections. However, the source of these formylated peptides is not clear, as most bacterial proteins are co-translationally deformylated by peptide deformylase. Here we develop a deformylation assay with translating ribosomes as substrates, to show that the binding of the signal recognition particle (SRP) to signal sequences in nascent proteins on the ribosome prevents deformylation, whereas deformylation of nascent proteins without signal sequence is not affected. Deformylation and its inhibition by SRP are not influenced by trigger factor, a chaperone that interacts with nascent chains on the ribosome. We propose that bacterial inner-membrane proteins, in particular those with N-out topology, can retain their N-terminal formyl group during cotranslational membrane insertion and supply formylated peptides during bacterial infections.

The conserved bacterial transcription terminator, Rho, is a potent target for bactericidal agents. Psu, a bacteriophage P4 capsid protein, is capable of inducing anti-termination to the Rho-dependent transcription termination. Knowledge of structural and mechanistic basis of this anti-termination is required to design peptide-inhibitor(s) of Rho from Psu. Using suppressor genetics, cross-linking, protein foot-printing and FRET analyses, we describe a conserved disordered structure, encompassing 139–153 amino acids of Rho, as the primary docking site for Psu. Also a neighbouring helical structure, comprising 347–354 amino acids, lining its central channel, plays a supportive role in the Rho–Psu complex formation. Based on the crystal structure of Psu, its conformation in the capsid of the P4 phage, and its interacting regions on Rho, we build an energy-minimized structural model of the Rho:Psu complex. In this model, a V-shaped dimer of Psu interacts with the two diagonally opposite subunits of a hexameric Rho, enabling Psu to form a ‘lid’ on the central channel of the latter. We show that this configuration of Psu makes the central channel of Rho inaccessible, and it causes a mechanical impediment to its translocase activity.

Background: Phage P4 Psu protein is a capsid decoration protein with unknown structure. Results: The first structure of Psu reveals a novel fold and a knotted dimer. Conclusion: The V-shaped molecular architecture is important for capsid binding. Significance: The structure of Psu will help to design peptide fragments, which can be used as drugs against the bacterial transcription machinery.

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