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Microbiology and Molecular Biology... Mar 2003Phage T4 has provided countless contributions to the paradigms of genetics and biochemistry. Its complete genome sequence of 168,903 bp encodes about 300 gene products.... (Review)
Review
Phage T4 has provided countless contributions to the paradigms of genetics and biochemistry. Its complete genome sequence of 168,903 bp encodes about 300 gene products. T4 biology and its genomic sequence provide the best-understood model for modern functional genomics and proteomics. Variations on gene expression, including overlapping genes, internal translation initiation, spliced genes, translational bypassing, and RNA processing, alert us to the caveats of purely computational methods. The T4 transcriptional pattern reflects its dependence on the host RNA polymerase and the use of phage-encoded proteins that sequentially modify RNA polymerase; transcriptional activator proteins, a phage sigma factor, anti-sigma, and sigma decoy proteins also act to specify early, middle, and late promoter recognition. Posttranscriptional controls by T4 provide excellent systems for the study of RNA-dependent processes, particularly at the structural level. The redundancy of DNA replication and recombination systems of T4 reveals how phage and other genomes are stably replicated and repaired in different environments, providing insight into genome evolution and adaptations to new hosts and growth environments. Moreover, genomic sequence analysis has provided new insights into tail fiber variation, lysis, gene duplications, and membrane localization of proteins, while high-resolution structural determination of the "cell-puncturing device," combined with the three-dimensional image reconstruction of the baseplate, has revealed the mechanism of penetration during infection. Despite these advances, nearly 130 potential T4 genes remain uncharacterized. Current phage-sequencing initiatives are now revealing the similarities and differences among members of the T4 family, including those that infect bacteria other than Escherichia coli. T4 functional genomics will aid in the interpretation of these newly sequenced T4-related genomes and in broadening our understanding of the complex evolution and ecology of phages-the most abundant and among the most ancient biological entities on Earth.
Topics: Bacteriophage T4; DNA Repair; DNA Replication; DNA, Viral; Genome, Viral; Lysogeny; Models, Molecular; Oligonucleotide Array Sequence Analysis; Promoter Regions, Genetic; Protein Biosynthesis; RNA, Viral; Ribosomes; Transcription, Genetic
PubMed: 12626685
DOI: 10.1128/MMBR.67.1.86-156.2003 -
Viruses Feb 2023Bacteriophage (phage) T4 has served as an extraordinary model to elucidate biological structures and mechanisms. Recent discoveries on the T4 head (capsid) structure,... (Review)
Review
Bacteriophage (phage) T4 has served as an extraordinary model to elucidate biological structures and mechanisms. Recent discoveries on the T4 head (capsid) structure, portal vertex, and genome packaging add a significant body of new literature to phage biology. Head structures in unexpanded and expanded conformations show dramatic domain movements, structural remodeling, and a ~70% increase in inner volume while creating high-affinity binding sites for the outer decoration proteins Soc and Hoc. Small changes in intercapsomer interactions modulate angles between capsomer planes, leading to profound alterations in head length. The in situ cryo-EM structure of the symmetry-mismatched portal vertex shows the remarkable structural morphing of local regions of the portal protein, allowing similar interactions with the capsid protein in different structural environments. Conformational changes in these interactions trigger the structural remodeling of capsid protein subunits surrounding the portal vertex, which propagate as a wave of expansion throughout the capsid. A second symmetry mismatch is created when a pentameric packaging motor assembles at the outer "clip" domains of the dodecameric portal vertex. The single-molecule dynamics of the packaging machine suggests a continuous burst mechanism in which the motor subunits adjusted to the shape of the DNA fire ATP hydrolysis, generating speeds as high as 2000 bp/s.
Topics: Bacteriophage T4; Binding Sites; Capsid; Capsid Proteins; Head
PubMed: 36851741
DOI: 10.3390/v15020527 -
Archives of Microbiology Jul 2014Analysis of molecular events in T4-infected Escherichia coli has revealed some of the most important principles of biology, including relationships between structures of... (Review)
Review
Analysis of molecular events in T4-infected Escherichia coli has revealed some of the most important principles of biology, including relationships between structures of genes and their products, virus-induced acquisition of metabolic function, and morphogenesis of complex structures through sequential gene product interaction rather than sequential gene activation. T4 bacteriophages and related strains were applied in the first formulations of many fundamental biological concepts. These include the unambiguous recognition of nucleic acids as the genetic material, the definition of the gene by fine-structure mutation, recombinational and functional analyses, the demonstration that the genetic code is triplet, the discovery of mRNA, the importance of recombination and DNA replications, light-dependent and light-independent DNA repair mechanisms, restriction and modification of DNA, self-splicing of intron/exon arrangement in prokaryotes, translation bypassing and others. Bacteriophage T4 possesses unique features that make it a good tool for a multicomponent vaccine platform. Hoc/Soc-fused antigens can be assembled on the T4 capsid in vitro and in vivo. T4-based phage display combined with affinity chromatography can be applied as a new method for bacteriophage purification. The T4 phage display system can also be used as an attractive approach for cancer therapy. The data show the efficient display of both single and multiple HIV antigens on the phage T4 capsid and offer insights for designing novel particulate HIV or other vaccines that have not been demonstrated by other vector systems.
Topics: Bacteriophage T4; Capsid; Cell Surface Display Techniques; Escherichia coli; Peptide Library; Vaccines
PubMed: 24828789
DOI: 10.1007/s00203-014-0989-8 -
Virology Journal Oct 2010
Topics: Bacteriophage T4
PubMed: 21029437
DOI: 10.1186/1743-422X-7-293 -
Viruses Mar 2022Viruses are biochemically complex structures and mainly consist of folded proteins that contain nucleic acids. Bacteriophage T4 is one of most prominent examples, having... (Review)
Review
Viruses are biochemically complex structures and mainly consist of folded proteins that contain nucleic acids. Bacteriophage T4 is one of most prominent examples, having a tail structure that contracts during the infection process. Intracellular phage multiplication leads to separate self-directed assembly reactions of proheads, tails and tail fibers. The proheads are packaged with concatemeric DNA produced by tandem replication reactions of the parental DNA molecule. Once DNA packaging is completed, the head is joined with the tail and six long fibers are attached. The mature particles are then released from the cell via lysis, another tightly regulated process. These processes have been studied in molecular detail leading to a fascinating view of the protein-folding dynamics that direct the structural interplay of assembled complexes. Lindsay W. Black dedicated his career to identifying and defining the molecular events required to form the T4 virion. He leaves us with rich insights into the astonishingly precise molecular clockwork that co-ordinates all of the players in T4 assembly, both viral and cellular. Here, we summarize Lindsay's key research contributions that are certain to stimulate our future science for many years to come.
Topics: Bacteriophage T4; Beauty; Capsid; DNA Packaging; DNA, Viral
PubMed: 35458430
DOI: 10.3390/v14040700 -
Future Microbiology 2014Bacteriophage T4 is the most well-studied member of Myoviridae, the most complex family of tailed phages. T4 assembly is divided into three independent pathways: the... (Review)
Review
Bacteriophage T4 is the most well-studied member of Myoviridae, the most complex family of tailed phages. T4 assembly is divided into three independent pathways: the head, the tail and the long tail fibers. The prolate head encapsidates a 172 kbp concatemeric dsDNA genome. The 925 Å-long tail is surrounded by the contractile sheath and ends with a hexagonal baseplate. Six long tail fibers are attached to the baseplate's periphery and are the host cell's recognition sensors. The sheath and the baseplate undergo large conformational changes during infection. X-ray crystallography and cryo-electron microscopy have provided structural information on protein-protein and protein-nucleic acid interactions that regulate conformational changes during assembly and infection of Escherichia coli cells.
Topics: Bacteriophage T4; Cryoelectron Microscopy; Crystallography, X-Ray; Escherichia coli; Genome, Viral; Models, Molecular; Protein Conformation; Protein Structure, Tertiary; Viral Proteins; Virus Assembly
PubMed: 25517898
DOI: 10.2217/fmb.14.91 -
Nature Communications May 2023Designing artificial viral vectors (AVVs) programmed with biomolecules that can enter human cells and carry out molecular repairs will have broad applications. Here, we...
Designing artificial viral vectors (AVVs) programmed with biomolecules that can enter human cells and carry out molecular repairs will have broad applications. Here, we describe an assembly-line approach to build AVVs by engineering the well-characterized structural components of bacteriophage T4. Starting with a 120 × 86 nm capsid shell that can accommodate 171-Kbp DNA and thousands of protein copies, various combinations of biomolecules, including DNAs, proteins, RNAs, and ribonucleoproteins, are externally and internally incorporated. The nanoparticles are then coated with cationic lipid to enable efficient entry into human cells. As proof of concept, we assemble a series of AVVs designed to deliver full-length dystrophin gene or perform various molecular operations to remodel human genome, including genome editing, gene recombination, gene replacement, gene expression, and gene silencing. These large capacity, customizable, multiplex, and all-in-one phage-based AVVs represent an additional category of nanomaterial that could potentially transform gene therapies and personalized medicine.
Topics: Humans; Bacteriophage T4; Genome, Human; Genetic Vectors; Capsid Proteins; Capsid; DNA, Viral
PubMed: 37253769
DOI: 10.1038/s41467-023-38364-1 -
Viruses Jun 2015The T4 bacteriophage encodes eight proteins, which are sufficient to carry out coordinated leading and lagging strand DNA synthesis. These purified proteins have been... (Review)
Review
The T4 bacteriophage encodes eight proteins, which are sufficient to carry out coordinated leading and lagging strand DNA synthesis. These purified proteins have been used to reconstitute DNA synthesis in vitro and are a well-characterized model system. Recent work on the T4 replisome has yielded more detailed insight into the dynamics and coordination of proteins at the replication fork. Since the leading and lagging strands are synthesized in opposite directions, coordination of DNA synthesis as well as priming and unwinding is accomplished by several protein complexes. These protein complexes serve to link catalytic activities and physically tether proteins to the replication fork. Essential to both leading and lagging strand synthesis is the formation of a holoenzyme complex composed of the polymerase and a processivity clamp. The two holoenzymes form a dimer allowing the lagging strand polymerase to be retained within the replisome after completion of each Okazaki fragment. The helicase and primase also form a complex known as the primosome, which unwinds the duplex DNA while also synthesizing primers on the lagging strand. Future studies will likely focus on defining the orientations and architecture of protein complexes at the replication fork.
Topics: Bacteriophage T4; DNA Replication; DNA, Viral; Multienzyme Complexes; Viral Proteins
PubMed: 26102578
DOI: 10.3390/v7062766 -
Virology Journal Dec 2010The bacteriophage T4 encodes 10 proteins, known collectively as the replisome, that are responsible for the replication of the phage genome. The replisomal proteins can... (Review)
Review
The bacteriophage T4 encodes 10 proteins, known collectively as the replisome, that are responsible for the replication of the phage genome. The replisomal proteins can be subdivided into three activities; the replicase, responsible for duplicating DNA, the primosomal proteins, responsible for unwinding and Okazaki fragment initiation, and the Okazaki repair proteins. The replicase includes the gp43 DNA polymerase, the gp45 processivity clamp, the gp44/62 clamp loader complex, and the gp32 single-stranded DNA binding protein. The primosomal proteins include the gp41 hexameric helicase, the gp61 primase, and the gp59 helicase loading protein. The RNaseH, a 5' to 3' exonuclease and T4 DNA ligase comprise the activities necessary for Okazaki repair. The T4 provides a model system for DNA replication. As a consequence, significant effort has been put forth to solve the crystallographic structures of these replisomal proteins. In this review, we discuss the structures that are available and provide comparison to related proteins when the T4 structures are unavailable. Three of the ten full-length T4 replisomal proteins have been determined; the gp59 helicase loading protein, the RNase H, and the gp45 processivity clamp. The core of T4 gp32 and two proteins from the T4 related phage RB69, the gp43 polymerase and the gp45 clamp are also solved. The T4 gp44/62 clamp loader has not been crystallized but a comparison to the E. coli gamma complex is provided. The structures of T4 gp41 helicase, gp61 primase, and T4 DNA ligase are unknown, structures from bacteriophage T7 proteins are discussed instead. To better understand the functionality of T4 DNA replication, in depth structural analysis will require complexes between proteins and DNA substrates. A DNA primer template bound by gp43 polymerase, a fork DNA substrate bound by RNase H, gp43 polymerase bound to gp32 protein, and RNase H bound to gp32 have been crystallographically determined. The preparation and crystallization of complexes is a significant challenge. We discuss alternate approaches, such as small angle X-ray and neutron scattering to generate molecular envelopes for modeling macromolecular assemblies.
Topics: Bacteriophage T4; Bacteriophage T7; Crystallography, X-Ray; DNA Replication; Macromolecular Substances; Models, Biological; Models, Molecular; Protein Structure, Quaternary; Protein Structure, Tertiary; Scattering, Small Angle; Viral Proteins
PubMed: 21129204
DOI: 10.1186/1743-422X-7-359 -
Biochemistry. Biokhimiia Nov 2004In studying bacteriophage T4--one of the basic models of molecular biology for several decades--there has come a Renaissance, and this virus is now actively used as... (Review)
Review
In studying bacteriophage T4--one of the basic models of molecular biology for several decades--there has come a Renaissance, and this virus is now actively used as object of structural biology. The structures of six proteins of the phage particle have recently been determined at atomic resolution by X-ray crystallography. Three-dimensional reconstruction of the infection device--one of the most complex multiprotein components--has been developed on the basis of cryo-electron microscopy images. The further study of bacteriophage T4 structure will allow a better understanding of the regulation of protein folding, assembly of biological structures, and also mechanisms of functioning of the complex biological molecular machines.
Topics: Animals; Bacteriophage T4; Protein Conformation; Viral Proteins; Virus Assembly
PubMed: 15627372
DOI: 10.1007/s10541-005-0064-9