Cryo-electron microscopy of the f1 filamentous phage reveals a new paradigm in viral infection and assembly

Abstract

Phages are viruses that infect bacteria and dominate every ecosystem on our planet. As well as impacting microbial ecology, physiology and evolution, phages are exploited as tools in molecular biology and biotechnology. This is particularly true for the Ff (f1, fd or M13) phages, which represent a widely distributed group of filamentous viruses. Over nearly five decades, Ff has seen an extraordinary range of applications, including in phage display and nanotechnology. However, the complete structure of the phage capsid and consequently the mechanisms of infection and assembly remain largely mysterious. Using cryo-electron microscopy and a highly efficient system for production of short Ff-derived nanorods, we have determined the first structure of a filamentous virus, including the filament tips. Structure combined with mutagenesis was employed to identify domains of the phage that are important in bacterial attack and for release of new phage progeny. These data allow new models to be proposed for the phage lifecycle and will undoubtedly enable the development of novel biotechnological applications.

A packaging signal-binding protein regulates the assembly checkpoint of integrative filamentous phages

“…The structures were illustrated based on AlphaFold or PDB as the following: PSB15 (Uniport: Q8P905), pIII (Uniport: Q3BSR6), pVI (Uniport: Q3BSR8), pVII (Uniport: Q3BSR4), pVIII (PDB:2IFO), thioredoxin (PDB: 2TRX), phage (PDB: 2IFO, 8B3P, 8B3O, 8IXK, 8IXL). Both ends of phage are adopted from f1 and M13 phage structures (Conners et al, 2023; Jia and Xiang, 2023).…”

“…PS is a cis -acting DNA element that distinguishes viral genomes from cellular DNAs or RNAs (Tate and Peterson, 1974; Shen et al, 1979; Webster et al, 1981; Dotto et al, 1981). During encapsidation, directionality of Ff particles matters as phage particles are structurally asymmetric at their leading and trailing end (Wickner and Killick, 1977; Conners et al, 2023; Jia and Xiang, 2023). At the leading end, pVII and pIX protein complex interact with the PS hairpin of pV-coated phage ssDNA (Russel and Model, 1989; Russel, 1993; Haase et al, 2022).…”

Diversification of Phage-Displayed Peptide Libraries with Noncanonical Amino Acid Mutagenesis and Chemical Modification

“…22 Through recent Cryo-EM studies, it was determined that the C-terminal domain is primarily composed of alpha helices for insertion within the virion (Figure 2D). 34 The Cterminal end of the protein also contains a transmembrane region that serves to anchor the protein in the host membrane during assembly. 35…”

Single-cell imaging of the lytic phage life cycle in bacteria

“…The vast majority of current methods used to study the kinetics of phage infections steps in bacteria rely on bulk culture approaches and omics analysis [7][8][9] , which lack the necessary single-cell resolution to analyse cell-to-cell heterogeneity, or on cryo-EM imaging for high-resolution investigation of structural aspects, which lacks time-resolved information to monitor the progress of individual infection steps and their interrelations 10,11 . In order to quantify and understand how the physiological diversity of host cells influences the infection dynamics of phages, a method is needed that can precisely monitor each stage of infection initiated by a single phage targeting a solitary living bacterium in a time-resolved fashion.…”

“…10 ml -1Doubling timeDrawn from a normal distribution with mean 20 min, standard deviation 2.5 min. Positive definite.Steric limitMaximum number of phages that can adsorb to a single bacterial cell: 100.…”