Dataset Information

Novel DNA packaging recognition in the unusual bacteriophage N15.

ABSTRACT: Phage lambda's cosB packaging recognition site is tripartite, consisting of 3 TerS binding sites, called R sequences. TerS binding to the critical R3 site positions the TerL endonuclease for nicking cosN to generate cohesive ends. The N15 cos (cos(N15)) is closely related to cos(?), but whereas the cosB(N15) subsite has R3, it lacks the R2 and R1 sites and the IHF binding site of cosB(?). A bioinformatic study of N15-like phages indicates that cosB(N15) also has an accessory, remote rR2 site, which is proposed to increase packaging efficiency, like R2 and R1 of lambda. N15 plus five prophages all have the rR2 sequence, which is located in the TerS-encoding 1 gene, approximately 200 bp distal to R3. An additional set of four highly related prophages, exemplified by Monarch, has R3 sequence, but also has R2 and R1 sequences characteristic of cosB-?. The DNA binding domain of TerS-N15 is a dimer.

SUBMITTER: Feiss M

PROVIDER: S-EPMC4461450 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature

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Novel DNA packaging recognition in the unusual bacteriophage N15.

Feiss Michael M Geyer Henriette H Klingberg Franco F Moreno Norma N Forystek Amanda A Maluf Nasib Karl NK Sippy Jean J

Virology 20150516

Phage lambda's cosB packaging recognition site is tripartite, consisting of 3 TerS binding sites, called R sequences. TerS binding to the critical R3 site positions the TerL endonuclease for nicking cosN to generate cohesive ends. The N15 cos (cos(N15)) is closely related to cos(λ), but whereas the cosB(N15) subsite has R3, it lacks the R2 and R1 sites and the IHF binding site of cosB(λ). A bioinformatic study of N15-like phages indicates that cosB(N15) also has an accessory, remote rR2 site, wh ...[more]

PMID: 25956737

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Project description:Like many viruses, bacteriophage phi X174 packages its DNA genome into a procapsid that is assembled from structural intermediates and scaffolding proteins. The procapsid contains the structural proteins F, G and H, as well as the scaffolding proteins B and D. Provirions are formed by packaging of DNA together with the small internal J proteins, while losing at least some of the B scaffolding proteins. Eventually, loss of the D scaffolding proteins and the remaining B proteins leads to the formation of mature virions.phi X174 108S 'procapsids' have been purified in milligram quantities by removing 114S (mature virion) and 70S (abortive capsid) particles from crude lysates by differential precipitation with polyethylene glycol. 132S 'provirions' were purified on sucrose gradients in the presence of EDTA. Cryo-electron microscopy (cryo-EM) was used to obtain reconstructions of procapsids and provirions. Although these are very similar to each other, their structures differ greatly from that of the virion. The F and G proteins, whose atomic structures in virions were previously determined from X-ray crystallography, were fitted into the cryo-EM reconstructions. This showed that the pentamer of G proteins on each five-fold vertex changes its conformation only slightly during DNA packaging and maturation, whereas major tertiary and quaternary structural changes occur in the F protein. The procapsids and provirions were found to contain 120 copies of the D protein arranged as tetramers on the two-fold axes. DNA might enter procapsids through one of the 30 A diameter holes on the icosahedral three-fold axes.Combining cryo-EM image reconstruction and X-ray crystallography has revealed the major conformational changes that can occur in viral assembly. The function of the scaffolding proteins may be, in part, to support weak interactions between the structural proteins in the procapsids and to cover surfaces that are subsequently required for subunit-subunit interaction in the virion. The structures presented here are, therefore, analogous to chaperone proteins complexed with folding intermediates of a substrate.

Dataset Information

Novel DNA packaging recognition in the unusual bacteriophage N15.

Publications

Novel DNA packaging recognition in the unusual bacteriophage N15.

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