Three-dimensional structure of the bacteriophage P22 tail machine

Tang, Liang; Marion, William; Cingolani, Gino; Prevelige, Peter E.; Johnson, John E.

doi:10.1038/sj.emboj.7600695

Cited by 77 publications

(105 citation statements)

References 56 publications

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“…5a). This overall feature resembles that seen in other isolated portals with negligible sequence similarity [16][17][18][19] . The azimuthal density distribution of Epsilon15 portal deviates noticeably from perfect 12-fold symmetry (Fig.…”

supporting

confidence: 56%

Structure of epsilon15 bacteriophage reveals genome organization and DNA packaging/injection apparatus

Jiang

Chang

Jakana³

et al. 2006

Nature

286

340

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Using single particle electron cryomicroscopy that does not impose icosahedral averaging, we determined the structure of the entire infectious Salmonella phage Epsilon15 1 , including both icosahedral and non-icosahedral components. At least three layers of condensed viral DNA were observed to pack in coaxial coils with local 25 Å hexagonal inter-strand spacing. At one of the fivefold vertices, a portal complex with twelve subunits replaces a capsid pentamer. A tail hub with six projecting trimeric tailspikes sits on the external face of the portal. Below the portal is a cylindrical protein core. An extended shaft of density fills the central channel of the protein core and the portal complex and appears to consist of about 90 nucleotides at the terminus of the packaged DNA poised for injection. Using an icosahedral symmetry imposed reconstruction, the fold of the capsid shell protein is seen to resemble the capsid protein fold of other tailed double-stranded DNA phages 2-5 and human herpesvirus 6 . These common structural features suggest a common evolutionary origin among these viruses. Double-stranded DNA (dsDNA) phages are vectors for gene transfer among enteric bacteria, including important human pathogens 7 . For all the well-studied tailed dsDNA phages, a preformed procapsid shell is assembled, and the DNA is pumped into the shell through a portal complex located at a single vertex 8 . The phage tails are also assembled at this vertex. The portal complex together with packaging enzymes have been shown to function as components of a very powerful molecular motor 9 , but it has not been possible to visualize the complex within the intact virion.The inability to visualize the packed DNA and the portal vertex in the virion reflects the difficulties in determining these structural features which lack icosahedral symmetry and are lost in any icosahedral averaging used in X-ray crystallography or electron cryomicroscopy (cryoEM). Using a cryoEM single particle reconstruction technique without symmetry imposition, we have been able to determine the structure of these critical features of Salmonella phage Epsilon15, some of which are unexpected.Epsilon15 is a short tailed dsDNA bacteriophage that infects Salmonella anatum. Its genome (NCBI accession number: NC_004775) contains 39,671 base pairs with 49 open reading frames ( Supplementary Fig. 1) among which six, coding for structural proteins, were resolved by SDS-PAGE and identified by tryptic-digest/mass spectrometry ( Supplementary Fig. 2).

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supporting

confidence: 56%

Structure of epsilon15 bacteriophage reveals genome organization and DNA packaging/injection apparatus

Jiang

Chang

Jakana³

et al. 2006

Nature

286

340

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“…The tail complex is a sophisticated molecular machine, which is attached to a unique vertex of the icosahedral capsid and provides an entry through which the viral genome is packaged during replication and is ejected into the host during infection (2). In the prototypical Salmonella enterica phage P22 (3), the tail machine consists of a ϳ2.8-MDa multisubunit complex (4,5) that replaces a single penton of the icosahedral capsid (6 -8). P22 binds to the Salmonella surface via tailspike proteins (gp9) that provide adsorption specificity by binding to the Salmonella O-antigen surface polysaccharide and by cleaving it (9).…”

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confidence: 99%

Structural Plasticity of the Protein Plug That Traps Newly Packaged Genomes in Podoviridae Virions

Bhardwaj

Sankhala

Olia

et al. 2016

Journal of Biological Chemistry

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Bacterial viruses of the P22-like family encode a specialized tail needle essential for genome stabilization after DNA packaging and implicated in Gram-negative cell envelope penetration. The atomic structure of P22 tail needle (gp26) crystallized at acidic pH reveals a slender fiber containing an N-terminal "trimer of hairpins" tip. Although the length and composition of tail needles vary significantly in Podoviridae, unexpectedly, the amino acid sequence of the N-terminal tip is exceptionally conserved in more than 200 genomes of P22-like phages and prophages. In this paper, we used x-ray crystallography and EM to investigate the neutral pH structure of three tail needles from bacteriophage P22, HK620, and Sf6. In all cases, we found that the N-terminal tip is poorly structured, in stark contrast to the compact trimer of hairpins seen in gp26 crystallized at acidic pH. Hydrogen-deuterium exchange mass spectrometry, limited proteolysis, circular dichroism spectroscopy, and gel filtration chromatography revealed that the N-terminal tip is highly dynamic in solution and unlikely to adopt a stable trimeric conformation at physiological pH. This is supported by the cryo-EM reconstruction of P22 mature virion tail, where the density of gp26 N-terminal tip is incompatible with a trimer of hairpins. We propose the tail needle N-terminal tip exists in two conformations: a pre-ejection extended conformation, which seals the portal vertex after genome packaging, and a postejection trimer of hairpins, which forms upon its release from the virion. The conformational plasticity of the tail needle N-terminal tip is built in the amino acid sequence, explaining its extraordinary conservation in nature.Podoviridae forms a family of bacterial viruses (bacteriophages) characterized by short and noncontractile tails (1). The tail complex is a sophisticated molecular machine, which is attached to a unique vertex of the icosahedral capsid and provides an entry through which the viral genome is packaged during replication and is ejected into the host during infection (2). In the prototypical Salmonella enterica phage P22 (3), the tail machine consists of a ϳ2.8-MDa multisubunit complex (4, 5) that replaces a single penton of the icosahedral capsid (6 -8). P22 binds to the Salmonella surface via tailspike proteins (gp9) that provide adsorption specificity by binding to the Salmonella O-antigen surface polysaccharide and by cleaving it (9). In addition to the tailspike, P22 virions contain a tail needle that is located at the distal tip of the tail axis (5-7), projecting ϳ140 Å outwards from the virion. In P22, the tail needle protein is encoded by gene 26 (10, 11) and at acid pH forms a 240-Å-long trimeric coiled-coil fiber containing three domains: an N-terminal tip (NTT), 3 a central ␣-helical coiled coil core, and a C-terminal tip (CTT) that folds as an inverted coiled-coil (12-14). Orthologues of gene 26 have been identified bioinformatically in hundreds of phage genomes and prophages (15). Significant sequence variability exists ...

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“…The tailspike head-binding domain (HBD) binds to the interface of the tail adaptor and the tail nozzle, and the tailspike receptorbinding domain (RBD) recognizes cell receptor and cleaves host O-antigen, pulling virions close to the host cell upon infection. Sequential assembly of tail components to the phage capsid has been recognized in several podoviruses such as T7 (16) and P22 (15,(17)(18)(19). This assembly mechanism differs from that in the other two families, Myoviridae and Siphoviridae, of which the phage head, tail, and tail fibers are assembled independently and subsequently join to form the complete virion (20).…”

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confidence: 99%

High-resolution structure of podovirus tail adaptor suggests repositioning of an octad motif that mediates the sequential tail assembly

Liang

Zhao

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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The sophisticated tail structures of DNA bacteriophages play essential roles in life cycles. Podoviruses P22 and Sf6 have short tails consisting of multiple proteins, among which is a tail adaptor protein that connects the portal protein to the other tail proteins. Assembly of the tail has been shown to occur in a sequential manner to ensure proper molecular interactions, but the underlying mechanism remains to be understood. Here, we report the high-resolution structure of the tail adaptor protein gp7 from phage Sf6. The structure exhibits distinct distribution of opposite charges on two sides of the molecule. A gp7 dodecameric ring model shows an entirely negatively charged surface, suggesting that the assembly of the dodecamer occurs through head-to-tail interactions of the bipolar monomers. The N-terminal helix-loop structure undergoes rearrangement compared with that of the P22 homolog complexed with the portal, which is achieved by repositioning of two consecutive repeats of a conserved octad sequence motif. We propose that the conformation of the N-terminal helix-loop observed in the Sf6-gp7 and P22 portal:gp4 complex represents the pre- and postassembly state, respectively. Such motif repositioning may serve as a conformational switch that creates the docking site for the tail nozzle only after the assembly of adaptor protein to the portal. In addition, the C-terminal portion of gp7 shows conformational flexibility, indicating an induced fit on binding to the portal. These results provide insight into the mechanistic role of the adaptor protein in mediating the sequential assembly of the phage tail.

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Three-dimensional structure of the bacteriophage P22 tail machine

Cited by 77 publications

References 56 publications

Structure of epsilon15 bacteriophage reveals genome organization and DNA packaging/injection apparatus

Structure of epsilon15 bacteriophage reveals genome organization and DNA packaging/injection apparatus

Structural Plasticity of the Protein Plug That Traps Newly Packaged Genomes in Podoviridae Virions

High-resolution structure of podovirus tail adaptor suggests repositioning of an octad motif that mediates the sequential tail assembly

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