scholarly journals Intravirion DNA Can Access the Space Occupied by the Bacteriophage P22 Ejection Proteins

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1504
Author(s):  
Justin C. Leavitt ◽  
Eddie B. Gilcrease ◽  
Brianna M. Woodbury ◽  
Carolyn M. Teschke ◽  
Sherwood R. Casjens
Keyword(s):  
Dna Packaging ◽  
Bacteriophage P22 ◽  
Portal Protein ◽  
Dna Molecule ◽  
Double Stranded Dna ◽  
Phage P22

Tailed double-stranded DNA bacteriophages inject some proteins with their dsDNA during infection. Phage P22 injects about 12, 12, and 30 molecules of the proteins encoded by genes 7, 16 and 20, respectively. After their ejection from the virion, they assemble into a trans-periplasmic conduit through which the DNA passes to enter the cytoplasm. The location of these proteins in the virion before injection is not well understood, although we recently showed they reside near the portal protein barrel in DNA-filled heads. In this report we show that when these proteins are missing from the virion, a longer than normal DNA molecule is encapsidated by the P22 headful DNA packaging machinery. Thus, the ejection proteins occupy positions within the virion that can be occupied by packaged DNA when they are absent.

scholarly journals The Generalized Transducing Salmonella Bacteriophage ES18: Complete Genome Sequence and DNA Packaging Strategy

2005 ◽  
Vol 187 (3) ◽  
pp. 1091-1104 ◽  
Author(s):  
Sherwood R. Casjens ◽  
Eddie B. Gilcrease ◽  
Danella A. Winn-Stapley ◽  
Petra Schicklmaier ◽  
Horst Schmieger ◽  
...  
Keyword(s):  
Shigella Flexneri ◽  
Bioinformatic Analysis ◽  
Dna Packaging ◽  
Phage Group ◽  
Double Stranded Dna ◽  
Functional Classes ◽  
Phage P22 ◽  
Genome Nucleotide Sequence ◽  
Group Members ◽  
Packaging Strategy

ABSTRACT The generalized transducing double-stranded DNA bacteriophage ES18 has an icosahedral head and a long noncontractile tail, and it infects both rough and smooth Salmonella enterica strains. We report here the complete 46,900-bp genome nucleotide sequence and provide an analysis of the sequence. Its 79 genes and their organization clearly show that ES18 is a member of the lambda-like (lambdoid) phage group; however, it contains a novel set of genes that program assembly of the virion head. Most of its integration-excision, immunity, Nin region, and lysis genes are nearly identical to those of the short-tailed Salmonella phage P22, while other early genes are nearly identical to Escherichia coli phages λ and HK97, S. enterica phage ST64T, or a Shigella flexneri prophage. Some of the ES18 late genes are novel, while others are most closely related to phages HK97, lambda, or N15. Thus, the ES18 genome is mosaically related to other lambdoid phages, as is typical for all group members. Analysis of virion DNA showed that it is circularly permuted and about 10% terminally redundant and that initiation of DNA packaging series occurs across an approximately 1-kbp region rather than at a precise location on the genome. This supports a model in which ES18 terminase can move substantial distances along the DNA between recognition and cleavage of DNA destined to be packaged. Bioinformatic analysis of large terminase subunits shows that the different functional classes of phage-encoded terminases can usually be predicted from their amino acid sequence.

scholarly journals Cryo-EM structure and in vitro DNA packaging of a thermophilic virus with supersized T=7 capsids

2019 ◽  
Vol 116 (9) ◽  
pp. 3556-3561 ◽  
Author(s):  
Oliver W. Bayfield ◽  
Evgeny Klimuk ◽  
Dennis C. Winkler ◽  
Emma L. Hesketh ◽  
Maria Chechik ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Structural Integrity ◽  
Thermus Thermophilus ◽  
Dna Packaging ◽  
Dna Viruses ◽  
Portal Protein ◽  
Single Vertex ◽  
Double Stranded Dna ◽  
Β Sheets

Double-stranded DNA viruses, including bacteriophages and herpesviruses, package their genomes into preformed capsids, using ATP-driven motors. Seeking to advance structural and mechanistic understanding, we established in vitro packaging for a thermostable bacteriophage, P23-45 of Thermus thermophilus. Both the unexpanded procapsid and the expanded mature capsid can package DNA in the presence of packaging ATPase over the 20 °C to 70 °C temperature range, with optimum activity at 50 °C to 65 °C. Cryo-EM reconstructions for the mature and immature capsids at 3.7-Å and 4.4-Å resolution, respectively, reveal conformational changes during capsid expansion. Capsomer interactions in the expanded capsid are reinforced by formation of intersubunit β-sheets with N-terminal segments of auxiliary protein trimers. Unexpectedly, the capsid has T=7 quasi-symmetry, despite the P23-45 genome being twice as large as those of known T=7 phages, in which the DNA is compacted to near-crystalline density. Our data explain this anomaly, showing how the canonical HK97 fold has adapted to double the volume of the capsid, while maintaining its structural integrity. Reconstructions of the procapsid and the expanded capsid defined the structure of the single vertex containing the portal protein. Together with a 1.95-Å resolution crystal structure of the portal protein and DNA packaging assays, these reconstructions indicate that capsid expansion affects the conformation of the portal protein, while still allowing DNA to be packaged. These observations suggest a mechanism by which structural events inside the capsid can be communicated to the outside.

scholarly journals Architect of Virus Assembly: the Portal Protein Nucleates Procapsid Assembly in Bacteriophage P22

2019 ◽  
Vol 93 (9) ◽  
Author(s):  
Tina Motwani ◽  
Carolyn M. Teschke
Keyword(s):  
Virus Assembly ◽  
Genetic Material ◽  
Scaffolding Protein ◽  
Scaffolding Proteins ◽  
Bacteriophage P22 ◽  
Particle Assembly ◽  
Portal Protein ◽  
Double Stranded Dna ◽  
Dsdna Viruses

ABSTRACTTailed double-stranded DNA (dsDNA) bacteriophages, herpesviruses, and adenoviruses package their genetic material into a precursor capsid through a dodecameric ring complex called the portal protein, which is located at a unique 5-fold vertex. In several phages and viruses, including T4, Φ29, and herpes simplex virus 1 (HSV-1), the portal forms a nucleation complex with scaffolding proteins (SPs) to initiate procapsid (PC) assembly, thereby ensuring incorporation of only one portal ring per capsid. However, for bacteriophage P22, the role of its portal protein in initiation of procapsid assembly is unclear. We have developed anin vitroP22 assembly assay where portal protein is coassembled into procapsid-like particles (PLPs). Scaffolding protein also catalyzes oligomerization of monomeric portal protein into dodecameric rings, possibly forming a scaffolding protein-portal protein nucleation complex that results in one portal ring per P22 procapsid. Here, we present evidence substantiating that the P22 portal protein, similarly to those of other dsDNA viruses, can act as an assembly nucleator. The presence of the P22 portal protein is shown to increase the rate of particle assembly and contribute to proper morphology of the assembled particles. Our results highlight a key function of portal protein as an assembly initiator, a feature that is likely conserved among these classes of dsDNA viruses.IMPORTANCEThe existence of a single portal ring is essential to the formation of infectious virions in the tailed double-stranded DNA (dsDNA) phages, herpesviruses, and adenoviruses and, as such, is a viable antiviral therapeutic target. How only one portal is selectively incorporated at a unique vertex is unclear. In many dsDNA viruses and phages, the portal protein acts as an assembly nucleator. However, early work on phage P22 assemblyin vivoindicated that the portal protein did not function as a nucleator for procapsid (PC) assembly, leading to the suggestion that P22 uses a unique mechanism for portal incorporation. Here, we show that portal protein nucleates assembly of P22 procapsid-like particles (PLPs). Addition of portal rings to an assembly reaction increases the rate of formation and yield of particles and corrects improper particle morphology. Our data suggest that procapsid assembly may universally initiate with a nucleation complex composed minimally of portal and scaffolding proteins (SPs).

Purification and organization of the gene 1 portal protein required for phage P22 DNA packaging

Biochemistry ◽  
1988 ◽  
Vol 27 (6) ◽  
pp. 1849-1856 ◽  
Author(s):  
Christopher Bazinet ◽  
Julyet Benbasat ◽  
Jonathan King ◽  
Jose Maria Carazo ◽  
Jose L. Carrascosa
Keyword(s):  
Dna Packaging ◽  
Portal Protein ◽  
Phage P22

scholarly journals Portal Protein: The Orchestrator of Capsid Assembly for the dsDNA Tailed Bacteriophages and Herpesviruses

2019 ◽  
Vol 6 (1) ◽  
pp. 141-160 ◽  
Author(s):  
Corynne L. Dedeo ◽  
Gino Cingolani ◽  
Carolyn M. Teschke
Keyword(s):  
Drug Target ◽  
Sequence Similarity ◽  
Antiviral Drug ◽  
Viral Assembly ◽  
Dna Packaging ◽  
Capsid Assembly ◽  
Portal Protein ◽  
Ring Assembly ◽  
Double Stranded Dna

Tailed, double-stranded DNA bacteriophages provide a well-characterized model system for the study of viral assembly, especially for herpesviruses and adenoviruses. A wealth of genetic, structural, and biochemical work has allowed for the development of assembly models and an understanding of the DNA packaging process. The portal complex is an essential player in all aspects of bacteriophage and herpesvirus assembly. Despite having low sequence similarity, portal structures across bacteriophages share the portal fold and maintain a conserved function. Due to their dynamic role, portal proteins are surprisingly plastic, and their conformations change for each stage of assembly. Because the maturation process is dependent on the portal protein, researchers have been working to validate this protein as a potential antiviral drug target. Here we review recent work on the role of portal complexes in capsid assembly, including DNA packaging, as well as portal ring assembly and incorporation and analysis of portal structures.

scholarly journals High-resolution mapping of heteroduplex DNA formed during UV-induced and spontaneous mitotic recombination events in yeast

2017 ◽  
Author(s):  
Yi Yin ◽  
Margaret Dominska ◽  
Eunice Yim ◽  
Thomas D. Petes
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Mitotic Recombination ◽  
Template Switching ◽  
Wild Type ◽  
Dna Breaks ◽  
Repair Synthesis ◽  
Heteroduplex Dna ◽  
Dna Molecule ◽  
Double Stranded Dna

AbstractDouble-stranded DNA breaks (DSBs) can be generated by both endogenous and exogenous agents. In diploid yeast strains, such breaks are usually repaired by homologous recombination (HR), and a number of different HR pathways have been described. An early step for all HR pathways is formation of a heteroduplex, in which a single-strand from the broken DNA molecule pairs with a strand derived from an intact DNA molecule. If the two strands of DNA are not identical, within the heteroduplex DNA (hetDNA), there will be mismatches. In a wild-type strain, these mismatches are removed by the mismatch repair (MMR) system. In strains lacking MMR, the mismatches persist and can be detected by a variety of genetic and physical techniques. Most previous studies involving hetDNA formed during mitotic recombination have been restricted to a single locus with DSBs induced at a defined position by a site-specific endonuclease. In addition, in most of these studies, recombination between repeated genes was examined; in such studies, the sequence homologies were usually less than 5 kb. In the present study, we present a global mapping of hetDNA formed in a UV-treated MMR-defective mlh1 strain. Although about two-thirds of the recombination events were associated with hetDNA with a continuous array of unrepaired mismatches, in about one-third of the events, we found regions of unrepaired mismatches flanking regions without mismatches. We suggest that these discontinuous hetDNAs involve template switching during repair synthesis, repair of a double-stranded DNA gap, and/or Mlh1-independent MMR. Many of our observed events are not explicable by the simplest form of the double-strand break repair (DSBR) model of recombination. We also studied hetDNA associated with spontaneous recombination events selected on chromosomes IV and V in a wild-type strain. The interval on chromosome IV contained a hotspot for spontaneous crossovers generated by an inverted pair of transposable elements (HS4). We showed that HS4-induced recombination events are associated with the formation of very large (>30 kb) double-stranded DNA gaps.

Exploration of Spin-Dependent Thermoelectricity in the Chiral Double-Stranded DNA Molecule Coupled to Ferromagnetic Leads

2019 ◽  
Vol 12 (2) ◽  
Author(s):  
Lei-Lei Nian ◽  
Long Bai ◽  
Wenting Yu ◽  
Jun Tang ◽  
Huichao Li ◽  
...  
Keyword(s):  
Dna Molecule ◽  
Double Stranded Dna
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