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Viruses Jun 2019In the rhizosphere, bacteria-phage interactions are likely to have important impacts on the ecology of microbial communities and microbe-plant interactions. To better...
In the rhizosphere, bacteria-phage interactions are likely to have important impacts on the ecology of microbial communities and microbe-plant interactions. To better understand the dynamics of Agrobacteria-phage interactions, we have isolated diverse bacteriophages which infect the bacterial plant pathogen, . Here, we complete the genomic characterization of phages Atu_ph04 and Atu_ph08. Atu_ph04-a T4-like phage belonging to the family-was isolated from waste water and has a 143,349 bp genome that encodes 223 predicted open reading frames (ORFs). Based on phylogenetic analysis and whole-genome alignments, Atu_ph04 is a member of a newly described T4 superfamily that contains other -infecting phages. Atu_ph08, a member of the T7-like family, was isolated from waste water, has a 59,034 bp genome, and encodes 75 ORFs. Based on phylogenetic analysis and whole-genome alignments, Atu_ph08 may form a new T7 superfamily which includes phage PCB5 and phage POI1126. Atu_ph08 is predicted to have lysogenic activity, as we found evidence of an integrase and several transcriptional repressors with similarity to proteins in transducing phage P22. Together, this data suggests that phages are diverse in morphology, genomic content, and lifestyle.
Topics: Agrobacterium tumefaciens; Bacteriophage T4; Bacteriophage T7; Biological Control Agents; DNA, Viral; Genes, Viral; Genome, Viral; Genomics; Host Specificity; Myoviridae; Open Reading Frames; Phylogeny; Plant Diseases; Podoviridae; Sequence Alignment; Sequence Analysis, DNA; Virion
PubMed: 31181591
DOI: 10.3390/v11060528 -
Journal of Bacteriology Nov 1994
Review
Topics: Amino Acid Sequence; Bacteriophage T4; Catalysis; Conserved Sequence; DNA Mutational Analysis; Muramidase; Structure-Activity Relationship
PubMed: 7961435
DOI: 10.1128/jb.176.22.6783-6788.1994 -
MicrobiologyOpen Dec 2016Bacteriophages have strict host specificity and the step of adsorption is one of key factors for determining host specificity. Here, we systematically examined the...
Bacteriophages have strict host specificity and the step of adsorption is one of key factors for determining host specificity. Here, we systematically examined the interaction between the Escherichia coli receptors lipopolysaccharide (LPS) and outer membrane protein C (OmpC), and the long tail fibers of bacteriophage T4. Using a variety of LPS mutants, we demonstrated that T4 has no specificity for the sugar sequence of the outer core (one of three LPS regions) in the presence of OmpC but, in the absence of OmpC, can adsorb to a specific LPS which has only one or two glucose residues without a branch. These results strengthen the idea that T4 adsorbs to E. coli via two distinct modes, OmpC-dependent and OmpC-independent, suggested by previous reports (Prehm et al. 1976; Yu and Mizushima 1982). Isolation and characterization of the T4 mutants Nik (No infection to K-12 strain), Nib (No infection to B strain), and Arl (altered recognition of LPS) identified amino acids of the long tail fiber that play important roles in the interaction with OmpC or LPS, suggesting that the top surface of the distal tip head domain of T4 long tail fibers interacts with LPS and its lateral surface interacts with OmpC.
Topics: Amino Acid Substitution; Bacterial Outer Membrane Proteins; Bacteriophage T4; Escherichia coli; Host Specificity; Lipopolysaccharides; Porins; Receptors, Virus; Viral Tail Proteins; Virus Attachment
PubMed: 27273222
DOI: 10.1002/mbo3.384 -
Methods in Molecular Biology (Clifton,... 2017Protein-based subunit vaccines represent a safer alternative to the whole pathogen in vaccine development. However, limitations of physiological instability and low...
Protein-based subunit vaccines represent a safer alternative to the whole pathogen in vaccine development. However, limitations of physiological instability and low immunogenicity of such vaccines demand an efficient delivery system to stimulate robust immune responses. The bacteriophage T4 capsid-based antigen delivery system can robustly elicit both humoral and cellular immune responses without any adjuvant. Therefore, it offers a strong promise as a novel antigen delivery system. Currently Bacillus anthracis, the causative agent of anthrax, is a serious biothreat agent and no FDA-approved anthrax vaccine is available for mass vaccination. Here, we describe a potential anthrax vaccine using a T4 capsid platform to display and deliver the 83 kDa protective antigen, PA, a key component of the anthrax toxin. This T4 vaccine platform might serve as a universal antigen delivery system that can be adapted to develop vaccines against any infectious disease.
Topics: Anthrax Vaccines; Antigens, Bacterial; Bacterial Toxins; Bacteriophage T4; Capsid; Immunization; Nanoparticles
PubMed: 28374254
DOI: 10.1007/978-1-4939-6869-5_15 -
Virology Journal Oct 2010Control of transcription is crucial for correct gene expression and orderly development. For many years, bacteriophage T4 has provided a simple model system to... (Review)
Review
Control of transcription is crucial for correct gene expression and orderly development. For many years, bacteriophage T4 has provided a simple model system to investigate mechanisms that regulate this process. Development of T4 requires the transcription of early, middle and late RNAs. Because T4 does not encode its own RNA polymerase, it must redirect the polymerase of its host, E. coli, to the correct class of genes at the correct time. T4 accomplishes this through the action of phage-encoded factors. Here I review recent studies investigating the transcription of T4 prereplicative genes, which are expressed as early and middle transcripts. Early RNAs are generated immediately after infection from T4 promoters that contain excellent recognition sequences for host polymerase. Consequently, the early promoters compete extremely well with host promoters for the available polymerase. T4 early promoter activity is further enhanced by the action of the T4 Alt protein, a component of the phage head that is injected into E. coli along with the phage DNA. Alt modifies Arg265 on one of the two α subunits of RNA polymerase. Although work with host promoters predicts that this modification should decrease promoter activity, transcription from some T4 early promoters increases when RNA polymerase is modified by Alt. Transcription of T4 middle genes begins about 1 minute after infection and proceeds by two pathways: 1) extension of early transcripts into downstream middle genes and 2) activation of T4 middle promoters through a process called sigma appropriation. In this activation, the T4 co-activator AsiA binds to Region 4 of σ⁷⁰, the specificity subunit of RNA polymerase. This binding dramatically remodels this portion of σ⁷⁰, which then allows the T4 activator MotA to also interact with σ⁷⁰. In addition, AsiA restructuring of σ⁷⁰ prevents Region 4 from forming its normal contacts with the -35 region of promoter DNA, which in turn allows MotA to interact with its DNA binding site, a MotA box, centered at the -30 region of middle promoter DNA. T4 sigma appropriation reveals how a specific domain within RNA polymerase can be remolded and then exploited to alter promoter specificity.
Topics: Bacteriophage T4; DNA Replication; Escherichia coli; Gene Expression Regulation, Viral; Host-Parasite Interactions; Promoter Regions, Genetic; RNA, Messenger; RNA, Viral; Transcription, Genetic; Virus Replication
PubMed: 21029433
DOI: 10.1186/1743-422X-7-289 -
Proceedings of the National Academy of... Sep 2015The first stages of productive bacteriophage infections of bacterial host cells require efficient adsorption to the cell surface followed by ejection of phage DNA into...
The first stages of productive bacteriophage infections of bacterial host cells require efficient adsorption to the cell surface followed by ejection of phage DNA into the host cytoplasm. To achieve this goal, a phage virion must undergo significant structural remodeling. For phage T4, the most obvious change is the contraction of its tail. Here, we use skinny E. coli minicells as a host, along with cryo-electron tomography and mutant phage virions, to visualize key structural intermediates during initiation of T4 infection. We show for the first time that most long tail fibers are folded back against the tail sheath until irreversible adsorption, a feature compatible with the virion randomly walking across the cell surface to find an optimal site for infection. Our data confirm that tail contraction is triggered by structural changes in the baseplate, as intermediates were found with remodeled baseplates and extended tails. After contraction, the tail tube penetrates the host cell periplasm, pausing while it degrades the peptidoglycan layer. Penetration into the host cytoplasm is accompanied by a dramatic local outward curvature of the cytoplasmic membrane as it fuses with the phage tail tip. The baseplate hub protein gp27 and/or the ejected tape measure protein gp29 likely form the transmembrane channel for viral DNA passage into the cell cytoplasm. Building on the wealth of prior biochemical and structural information, this work provides new molecular insights into the mechanistic pathway of T4 phage infection.
Topics: Bacteriophage T4; Cell Membrane; Cryoelectron Microscopy; Genes, Viral; Host-Pathogen Interactions; Viral Tail Proteins; Virion
PubMed: 26283379
DOI: 10.1073/pnas.1501064112 -
Nature Communications Apr 2020Large biological structures are assembled from smaller, often symmetric, sub-structures. However, asymmetry among sub-structures is fundamentally important for...
Large biological structures are assembled from smaller, often symmetric, sub-structures. However, asymmetry among sub-structures is fundamentally important for biological function. An extreme form of asymmetry, a 12-fold-symmetric dodecameric portal complex inserted into a 5-fold-symmetric capsid vertex, is found in numerous icosahedral viruses, including tailed bacteriophages, herpesviruses, and archaeal viruses. This vertex is critical for driving capsid assembly, DNA packaging, tail attachment, and genome ejection. Here, we report the near-atomic in situ structure of the symmetry-mismatched portal vertex from bacteriophage T4. Remarkably, the local structure of portal morphs to compensate for symmetry-mismatch, forming similar interactions in different capsid environments while maintaining strict symmetry in the rest of the structure. This creates a unique and unusually dynamic symmetry-mismatched vertex that is central to building an infectious virion.
Topics: Bacteriophage T4; Capsid; Capsid Proteins; Cryoelectron Microscopy; DNA Packaging; DNA, Viral; Escherichia coli; Models, Molecular; Mutation; Viral Proteins; Virion; Virus Assembly
PubMed: 32249784
DOI: 10.1038/s41467-020-15575-4 -
Genetics Apr 1998Antimutators are mutant strains that have reduced mutation rates compared to the corresponding wild-type strain. Their existence, along with mutator mutants that have... (Review)
Review
Antimutators are mutant strains that have reduced mutation rates compared to the corresponding wild-type strain. Their existence, along with mutator mutants that have higher mutation rates compared to the wild-type strain, are powerful evidence that mutation rates are genetically controlled. Compared to mutator mutants, antimutators have a very distinguishing property. Because they prevent normally occurring mutations, they, uniquely, are capable of providing insight into the mechanisms of spontaneous mutations. In this review, antimutator mutants are discussed in bacteriophage T4 and the bacterium Escherichia coli, with regard to their properties, possible mechanisms, and implications for the sources of spontaneous mutations in these two organisms.
Topics: Bacteriophage T4; Escherichia coli; Mutation
PubMed: 9560377
DOI: 10.1093/genetics/148.4.1579 -
Proceedings of the National Academy of... Oct 2019Although many proteins possess a distinct folded structure lying at a minimum in a funneled free energy landscape, thermal energy causes any protein to continuously...
Although many proteins possess a distinct folded structure lying at a minimum in a funneled free energy landscape, thermal energy causes any protein to continuously access lowly populated excited states. The existence of excited states is an integral part of biological function. Although transitions into the excited states may lead to protein misfolding and aggregation, little structural information is currently available for them. Here, we show how NMR spectroscopy, coupled with pressure perturbation, brings these elusive species to light. As pressure acts to favor states with lower partial molar volume, NMR follows the ensuing change in the equilibrium spectroscopically, with residue-specific resolution. For T4 lysozyme L99A, relaxation dispersion NMR was used to follow the increase in population of a previously identified "invisible" folded state with pressure, as this is driven by the reduction in cavity volume by the flipping-in of a surface aromatic group. Furthermore, multiple partly disordered excited states were detected at equilibrium using pressure-dependent H/D exchange NMR spectroscopy. Here, unfolding reduced partial molar volume by the removal of empty internal cavities and packing imperfections through subglobal and global unfolding. A close correspondence was found for the distinct pressure sensitivities of various parts of the protein and the amount of internal cavity volume that was lost in each unfolding event. The free energies and populations of excited states allowed us to determine the energetic penalty of empty internal protein cavities to be 36 cal⋅Å.
Topics: Bacteriophage T4; Muramidase; Nuclear Magnetic Resonance, Biomolecular; Pressure; Protein Conformation; Protein Denaturation; Protein Folding; Proteins
PubMed: 31570587
DOI: 10.1073/pnas.1911181116 -
The Journal of Biological Chemistry Sep 2015Rad50 and Mre11 form a complex involved in the detection and processing of DNA double strand breaks. Rad50 contains an anti-parallel coiled-coil with two absolutely...
Rad50 and Mre11 form a complex involved in the detection and processing of DNA double strand breaks. Rad50 contains an anti-parallel coiled-coil with two absolutely conserved cysteine residues at its apex. These cysteine residues serve as a dimerization domain and bind a Zn(2+) cation in a tetrathiolate coordination complex known as the zinc-hook. Mutation of the zinc-hook in bacteriophage T4 is lethal, indicating the ability to bind Zn(2+) is critical for the functioning of the MR complex. In vitro, we found that complex formation between Rad50 and a peptide corresponding to the C-terminal domain of Mre11 enhances the ATPase activity of Rad50, supporting the hypothesis that the coiled-coil is a major conduit for communication between Mre11 and Rad50. We constructed mutations to perturb this domain in the bacteriophage T4 Rad50 homolog. Deletion of the Rad50 coiled-coil and zinc-hook eliminates Mre11 binding and ATPase activation but does not affect its basal activity. Mutation of the zinc-hook or disruption of the coiled-coil does not affect Mre11 or DNA binding, but their activation of Rad50 ATPase activity is abolished. Although these mutants excise a single nucleotide at a normal rate, they lack processivity and have reduced repetitive exonuclease rates. Restricting the mobility of the coiled-coil eliminates ATPase activation and repetitive exonuclease activity, but the ability to support single nucleotide excision is retained. These results suggest that the coiled-coiled domain adopts at least two conformations throughout the ATPase/nuclease cycle, with one conformation supporting enhanced ATPase activity and processivity and the other supporting nucleotide excision.
Topics: Adenosine Triphosphatases; Bacteriophage T4; DNA-Binding Proteins; Exonucleases; Mutation; Protein Structure, Tertiary; Viral Proteins; Zinc
PubMed: 26242734
DOI: 10.1074/jbc.M115.675132