Three-dimensional Structure of the Toxin-delivery Particle Antifeeding Prophage of Serratia entomophila

• Published in: The Journal of biological chemistry Vol. 288; no. 35; pp. 25276 - 25284
• Main Authors: Heymann, J. Bernard, Bartho, Joseph D., Rybakova, Daria, Venugopal, Hari P., Winkler, Dennis C., Sen, Anindito, Hurst, Mark R.H., Mitra, Alok K.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 30.08.2013; American Society for Biochemistry and Molecular Biology
• Subjects: Bacterial Pathogenesis; Bacterial Secretion Systems - physiology; Bacteriophage Tail Sheath; Bacteriophages - metabolism; Bacteriophages - ultrastructure; Cryo-electron Microscopy; Electron Microscopy (EM); Electron Tomography; Negative Stain Tomography; Protein Structure and Folding; Serratia - metabolism; Serratia - virology; Single Particle Analysis; Structural Biology; Bacteriophage Tail Sheath; Electron Tomography; Structural Biology; Bacterial Pathogenesis; Single Particle Analysis; Negative Stain Tomography; Electron Microscopy (EM); Cryo-electron Microscopy
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M113.456145

The Serratia entomophila antifeeding prophage (Afp) is a bullet-shaped toxin-delivery apparatus similar to the R-pyocins of Pseudomonas aeruginosa. Morphologically it resembles the sheathed tail of bacteriophages such as T4, including a baseplate at one end. It also shares features with the type VI secretion systems. Cryo-electron micrographs of tilted Afp specimens (up to 60 degrees) were analyzed to determine the correct cyclic symmetry to overcome the limitation imposed by exclusively side views in nominally untilted specimens. An asymmetric reconstruction shows clear 6-fold cyclic symmetry contrary to a previous conclusion of 4-fold symmetry based on analysis of only the preferred side views (Sen, A., Rybakova, D., Hurst, M. R., and Mitra, A. K. (2010) J. Bacteriol. 192, 4522–4525). Electron tomography of negatively stained Afp revealed right-handed helical striations in many of the particles, establishing the correct hand. Higher quality micrographs of untilted specimens were processed to produce a reconstruction at 2.0-nm resolution with imposed 6-fold symmetry. The helical parameters of the sheath were determined to be 8.14 nm for the subunit rise along and 40.5° for the rotation angle around the helix. The sheath is similar to that of the T4 phage tail but with a different arrangement of the subdomain of the polymerizing sheath protein(s). The central tube is similar to the diameter and axial width of the Hcp1 hexamer of P. aeruginosa type VI secretion system. The tube extends through the baseplate into a needle resembling the “puncture device” of the T4 tail. The tube contains density that may be the toxin and/or a length-determining protein. Background: Antifeeding prophage (Afp) is a toxin-delivery bacteriophage tail-like particle. Results: The syringe-like three-dimensional structure, composed of a helical sheath formed by 10 disks, a baseplate, and a central tube displays 6-fold symmetry. Conclusion: Although similar to other type VI secretion systems, Afp possesses unique features. Significance: This is the first insight into the three-dimensional structure of a tailocin.