A Defined in Vitro System for DNA Packaging by the Bacteriophage SPP1: Insights into the Headful Packaging Mechanism

Oliveira, Leonor; Alonso, Juan C.; Tavares, Paulo

doi:10.1016/j.jmb.2005.08.063

Cited by 40 publications

(59 citation statements)

References 47 publications

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“…Thus, we have to conclude that the right end of the genome and the length of the terminal redundancy remain unresolved, but it appears to be Ͼ8 kb. Furthermore, the findings suggest that the phage DNA may not be packed by the conventional pac-type headful mechanism (31). On the other hand, we estimated earlier that the phage genome size was 270 kb (14), which is 103% of the full-length genomic sequence reported here.…”

Section: Discussionmentioning

confidence: 51%

Characterization of the Genome, Proteome, and Structure of Yersiniophage ϕR1-37

Skurnik

Hyytiäinen

Happonen

et al. 2012

J Virol

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The bacteriophage vB_YecM-R1-37 (R1-37) is a lytic yersiniophage that can propagate naturally in different Yersinia species carrying the correct lipopolysaccharide receptor. This large-tailed phage has deoxyuridine (dU) instead of thymidine in its DNA. In this study, we determined the genomic sequence of phage R1-37, mapped parts of the phage transcriptome, characterized the phage particle proteome, and characterized the virion structure by cryo-electron microscopy and image reconstruction. The 262,391-bp genome of R1-37 is one of the largest sequenced phage genomes, and it contains 367 putative open reading frames (ORFs) and 5 tRNA genes. Mass-spectrometric analysis identified 69 phage particle structural proteins with the genes scattered throughout the genome. A total of 269 of the ORFs (73%) lack homologues in sequence databases. Based on terminator and promoter sequences identified from the intergenic regions, the phage genome was predicted to consist of 40 to 60 transcriptional units. Image reconstruction revealed that the R1-37 capsid consists of hexameric capsomers arranged on a T‫72؍‬ lattice similar to the bacteriophage KZ. The tail of R1-37 has a contractile sheath. We conclude that phage R1-37 is a representative of a novel phage type that carries the dU-containing genome in a KZ-like head. Bacteriophages, the viruses that infect bacteria, are the most abundant organisms on Earth, and it is estimated that for each microbial isolate at least 10 different phages exist (19,35). Present knowledge indicates that phages are extremely diverse in nature (7a). Studies on bacteriophages have escalated, since they are excellent targets for genomic and evolutionary research and as models for systems biology studies; in addition, they are important vehicles in horizontal gene transfer. Phages are used as tools in bacterial genetics, and phage gene products are used as tools in molecular biology. Furthermore, their potential as therapeutic agents during the increasing emergence of antibiotic resistance is being reexamined (45,46). In summary, a thorough knowledge of the bacteriophage and its biology is considered essential to all phage research.We have isolated several Yersinia enterocolitica-specific bacteriophages that use different parts of lipopolysaccharide (LPS) as receptors and used them to study the molecular biology and genetics of LPS biosynthesis (2,23,24,33,34,(46)(47)(48). The bacteriophage vB_YecM-R1-37 (R1-37) was isolated from sewage based on its ability to infect Y. enterocolitica strain YeO3-R1, an O-polysaccharide (O-PS)-lacking Y. enterocolitica serotype O:3 strain (44, 48). The host range of R1-37, as well as genetic and structural data, showed that the LPS outer core (OC) hexasaccharide of Y. enterocolitica O:3 is the phage receptor (23,37,38,48).Electron microscopy and analysis of its genome indicated that R1-37 is an exceptionally large-tailed phage with an estimated genome size of 270 kb (23). Structural studies on large bacteriophages with contractile tails are currently limited to the Pse...

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Section: Discussionmentioning

confidence: 51%

Characterization of the Genome, Proteome, and Structure of Yersiniophage ϕR1-37

Skurnik

Hyytiäinen

Happonen

et al. 2012

J Virol

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“…Samples were then mixed with BL21 E. coli bacteria, plated on soft agar, and incubated (overnight, 37°C) to determine viral titer. DNase protection assays were performed as described (26). For the reactions, the same T7/LPS ratio was incubated (37°C) for the times indicated above.…”

Section: Methodsmentioning

confidence: 99%

Conformational Changes Leading to T7 DNA Delivery upon Interaction with the Bacterial Receptor

González-García

Pulido-Cid

Garcia-Doval

et al. 2015

Journal of Biological Chemistry

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Background: T7 bacteriophage infects E. coli bacteria; during this process, the tail recognizes the bacterial receptor. Results: Interactions of T7 with rough LPS trigger DNA delivery by promoting changes in the tail tube. Conclusion: E. coli rough LPS act as the receptor in vitro for T7 bacteriophage. Significance: Biotechnological application of bacteriophages as antibacterial agents demands detailed molecular knowledge of bacteriophage infection.

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“…This is followed by the assembly of the DNA-packaging motor at the portal vertex of the shell. The motor pumps the SPP1 dsDNA into the capsid (8,39). It consists of gp6, gp1 (SPP1 DNA binding protein), and gp2 (viral ATPase/endonuclease).…”

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confidence: 99%

Capsid Structure and Its Stability at the Late Stages of Bacteriophage SPP1 Assembly

White

Sherman

Brasilès

et al. 2012

J Virol

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eThe structure of the bacteriophage SPP1 capsid was determined at subnanometer resolution by cryo-electron microscopy and single-particle analysis. The icosahedral capsid is composed of the major capsid protein gp13 and the auxiliary protein gp12, which are organized in a T‫7؍‬ lattice. DNA is arranged in layers with a distance of ϳ24.5 Å. gp12 forms spikes that are anchored at the center of gp13 hexamers. In a gp12-deficient mutant, the centers of hexamers are closed by loops of gp13 coming together to protect the SPP1 genome from the outside environment. The HK97-like fold was used to build a pseudoatomic model of gp13. Its structural organization remains unchanged upon tail binding and following DNA release. gp13 exhibits enhanced thermostability in the DNA-filled capsid. A remarkable convergence between the thermostability of the capsid and those of the other virion components was found, revealing that the overall architecture of the SPP1 infectious particle coevolved toward high robustness.

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A Defined in Vitro System for DNA Packaging by the Bacteriophage SPP1: Insights into the Headful Packaging Mechanism

Cited by 40 publications

References 47 publications

Characterization of the Genome, Proteome, and Structure of Yersiniophage ϕR1-37

Characterization of the Genome, Proteome, and Structure of Yersiniophage ϕR1-37

Conformational Changes Leading to T7 DNA Delivery upon Interaction with the Bacterial Receptor

Capsid Structure and Its Stability at the Late Stages of Bacteriophage SPP1 Assembly

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