Browse
Submit Data
Databases
API
Help

Dataset Information

25 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Bacteriophage T5 gene D10 encodes a branch-migration protein.

ABSTRACT: Helicases catalyze the unwinding of double-stranded nucleic acids where structure and phosphate backbone contacts, rather than nucleobase sequence, usually determines substrate specificity. We have expressed and purified a putative helicase encoded by the D10 gene of bacteriophage T5. Here we report that this hitherto uncharacterized protein possesses branch migration and DNA unwinding activity. The initiation of substrate unwinding showed some sequence dependency, while DNA binding and DNA-dependent ATPase activity did not. DNA footprinting and purine-base interference assays demonstrated that D10 engages these substrates with a defined polarity that may be established by protein-nucleobase contacts. Bioinformatic analysis of the nucleotide databases revealed genes predicted to encode proteins related to D10 in archaebacteria, bacteriophages and in viruses known to infect a range of eukaryotic organisms.

SUBMITTER: Wong IN

PROVIDER: S-EPMC5180179 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Bacteriophage T5 gene D10 encodes a branch-migration protein.

Wong Io Nam IN Sayers Jon R JR Sanders Cyril M CM

Scientific reports 20161223

Helicases catalyze the unwinding of double-stranded nucleic acids where structure and phosphate backbone contacts, rather than nucleobase sequence, usually determines substrate specificity. We have expressed and purified a putative helicase encoded by the D10 gene of bacteriophage T5. Here we report that this hitherto uncharacterized protein possesses branch migration and DNA unwinding activity. The initiation of substrate unwinding showed some sequence dependency, while DNA binding and DNA-depe ...[more]

PMID: 28009009

Similar Datasets

Identification of the ssDNA-binding protein of bacteriophage T5: Implications for T5 replication.

Project description:In a recent study, we identified and characterized the long-elusive replicative single-stranded DNA-binding protein of bacteriophage T5, which we showed is related to the eukaryotic transcription coactivator PC4. Here, we provide an extended discussion of these data, report several additional observations and consider implications for the recombination-dependent replication mechanism of the T5 genus, which is still poorly understood.

| S-EPMC3897522 | biostudies-literature

Novel Escherichia coli RNA Polymerase Binding Protein Encoded by Bacteriophage T5.

Project description:The Escherichia coli bacteriophage T5 has three temporal classes of genes (pre-early, early, and late). All three classes are transcribed by host RNA polymerase (RNAP) containing the ?70 promoter specificity subunit. Molecular mechanisms responsible for the switching of viral transcription from one class to another remain unknown. Here, we find the product of T5 gene 026 (gpT5.026) in RNAP preparations purified from T5-infected cells and demonstrate in vitro its tight binding to E. coli RNAP. While proteins homologous to gpT5.026 are encoded by all T5-related phages, no similarities to proteins with known functions can be detected. GpT5.026 binds to two regions of the RNAP ? subunit and moderately inhibits RNAP interaction with the discriminator region of ?70-dependent promoters. A T5 mutant with disrupted gene 026 is viable, but the host cell lysis phase is prolongated and fewer virus particles are produced. During the mutant phage infection, the number of early transcripts increases, whereas the number of late transcripts decreases. We propose that gpT5.026 is part of the regulatory cascade that orchestrates a switch from early to late bacteriophage T5 transcription.

| S-EPMC7472727 | biostudies-literature

Purified tails from bacteriophage T5

Project description: Not available

| EMPIAR-11319 | biostudies-other

Cloning, sequencing, and recombinational analysis with bacteriophage BF23 of the bacteriophage T5 oad gene encoding the receptor-binding protein.

Project description:Binding of bacteriophage T5 to its receptor, the Escherichia coli FhuA protein, is mediated by tail protein pb5. In this article we confirm that pb5 is encoded by the T5 oad gene and describe the isolation, expression, and sequencing of this gene. In order to locate oad precisely, we analyzed recombinants between BF23, a T5-related phage with a different host range, and plasmid clones containing segments of the T5 chromosome. This analysis also showed that oad has little or no homology with hrs, the analogous BF23 gene. We were able to overproduce a protein that comigrates with pb5 after fusing a 2-kb segment containing oad to a phage T7 promoter. This segment contains an open reading frame that can encode a protein of the appropriate size. Its deduced amino acid sequence does not closely resemble that of any other protein in the database. The sequence upstream of the open reading frame shows typical characteristics of a promoter region with two overlapping, divergently orientated promoters.

| S-EPMC207253 | biostudies-other

High affinity anchoring of the decoration protein pb10 onto the bacteriophage T5 capsid.

Project description:Bacteriophage capsids constitute icosahedral shells of exceptional stability that protect the viral genome. Many capsids display on their surface decoration proteins whose structure and function remain largely unknown. The decoration protein pb10 of phage T5 binds at the centre of the 120 hexamers formed by the major capsid protein. Here we determined the 3D structure of pb10 and investigated its capsid-binding properties using NMR, SAXS, cryoEM and SPR. Pb10 consists of an α-helical capsid-binding domain and an Ig-like domain exposed to the solvent. It binds to the T5 capsid with a remarkably high affinity and its binding kinetics is characterized by a very slow dissociation rate. We propose that the conformational exchange events observed in the capsid-binding domain enable rearrangements upon binding that contribute to the quasi-irreversibility of the pb10-capsid interaction. Moreover we show that pb10 binding is a highly cooperative process, which favours immediate rebinding of newly dissociated pb10 to the 120 hexamers of the capsid protein. In extreme conditions, pb10 protects the phage from releasing its genome. We conclude that pb10 may function to reinforce the capsid thus favouring phage survival in harsh environments.

| S-EPMC5292684 | biostudies-literature

Insights into bacteriophage T5 structure from analysis of its morphogenesis genes and protein components.

Project description:Bacteriophage T5 represents a large family of lytic Siphoviridae infecting Gram-negative bacteria. The low-resolution structure of T5 showed the T=13 geometry of the capsid and the unusual trimeric organization of the tail tube, and the assembly pathway of the capsid was established. Although major structural proteins of T5 have been identified in these studies, most of the genes encoding the morphogenesis proteins remained to be identified. Here, we combine a proteomic analysis of T5 particles with a bioinformatic study and electron microscopic immunolocalization to assign function to the genes encoding the structural proteins, the packaging proteins, and other nonstructural components required for T5 assembly. A head maturation protease that likely accounts for the cleavage of the different capsid proteins is identified. Two other proteins involved in capsid maturation add originality to the T5 capsid assembly mechanism: the single head-to-tail joining protein, which closes the T5 capsid after DNA packaging, and the nicking endonuclease responsible for the single-strand interruptions in the T5 genome. We localize most of the tail proteins that were hitherto uncharacterized and provide a detailed description of the tail tip composition. Our findings highlight novel variations of viral assembly strategies and of virion particle architecture. They further recommend T5 for exploring phage structure and assembly and for deciphering conformational rearrangements that accompany DNA transfer from the capsid to the host cytoplasm.

| S-EPMC3911642 | biostudies-literature

Pre-early functions of bacteriophage T5 and its relatives.

Project description:Coliphage T5 injects its DNA in 2 steps: the first step transfer (FST) region 7.9% is injected and its genes are expressed and only then does the remainder (second step transfer, SST) of its DNA enter the cell. In the FST region, only 2 essential genes (A1 and A2) have been identified and a third (dmp) non-essential gene codes for a deoxyribonucleotide 5' monophosphatase. Thirteen additional putative ORFs are present in the FST region. Numerous properties have been attributed to FST region, including SST, host DNA degradation, inhibition of host RNA and protein synthesis, restriction insensitivity and protection of T5 DNA. These effects do not occur following infection with an A1 mutant. The A2 gene seems only to be involved in SST transfer. This is puzzling since there are more seemingly unrelated effects than there are essential genes to accomplish them and it is possible that some important genes were not identified. This review attempts to analyze these problems that were first identified in the 1970-80 s. In particular, an attempt is made to determine which potential ORFs are conserved in evolution (and thus likely to be important); by comparing T5 to 10 newly isolated and completely sequenced T5-like phages. A similar approach is used to identify conserved repeats, inverted repeats and palindromes that occur in all T5-like phages in the region containing the injection stop signal (iss) and the terminase substrate. Finally, an attempt is made to re-analyze the mechanism whereby T5 protects itself from the enzymes that degrade host DNA, from the RecBCD nuclease and from restriction enzymes. For all of these FST effects new hypotheses and possible new genetic and biochemical approaches are envisaged.

| S-EPMC4743489 | biostudies-literature

Branch-specific plasticity of a bifunctional dopamine circuit encodes protein hunger.

Project description:Free-living animals must not only regulate the amount of food they consume but also choose which types of food to ingest. The shifting of food preference driven by nutrient-specific hunger can be essential for survival, yet little is known about the underlying mechanisms. We identified a dopamine circuit that encodes protein-specific hunger in Drosophila The activity of these neurons increased after substantial protein deprivation. Activation of this circuit simultaneously promoted protein intake and restricted sugar consumption, via signaling to distinct downstream neurons. Protein starvation triggered branch-specific plastic changes in these dopaminergic neurons, thus enabling sustained protein consumption. These studies reveal a crucial circuit mechanism by which animals adjust their dietary strategy to maintain protein homeostasis.

| S-EPMC5513152 | biostudies-literature

Characterization of an unusual bipolar helicase encoded by bacteriophage T5.

Project description:Bacteriophage T5 has a 120 kb double-stranded linear DNA genome encoding most of the genes required for its own replication. This lytic bacteriophage has a burst size of ?500 new phage particles per infected cell, demonstrating that it is able to turn each infected bacterium into a highly efficient DNA manufacturing machine. To begin to understand DNA replication in this prodigious bacteriophage, we have characterized a putative helicase encoded by gene D2. We show that bacteriophage T5 D2 protein is the first viral helicase to be described with bipolar DNA unwinding activities that require the same core catalytic residues for unwinding in either direction. However, unwinding of partially single- and double-stranded DNA test substrates in the 3'-5' direction is more robust and can be distinguished from the 5'-3' activity by a number of features including helicase complex stability, salt sensitivity and the length of single-stranded DNA overhang required for initiation of helicase action. The presence of D2 in an early gene cluster, the identification of a putative helix-turn-helix DNA-binding motif outside the helicase core and homology with known eukaryotic and prokaryotic replication initiators suggest an involvement for this unusual helicase in DNA replication initiation.

| S-EPMC3632103 | biostudies-literature

Rad51 protein stimulates the branch migration activity of Rad54 protein.

Project description:The Rad51 and Rad54 proteins play important roles during homologous recombination in eukaryotes. Rad51 forms a nucleoprotein filament on single-stranded DNA and performs the initial steps of double strand break repair. Rad54 belongs to the Swi2/Snf2 family of ATP-dependent DNA translocases. We previously showed that Rad54 promotes branch migration of Holliday junctions. Here we find that human Rad51 (hRad51) significantly stimulates the branch migration activity of hRad54. The stimulation appears to be evolutionarily conserved, as yeast Rad51 also stimulates the branch migration activity of yeast Rad54. We further investigated the mechanism of this stimulation. Our results demonstrate that the stimulation of hRad54-promoted branch migration by hRad51 is driven by specific protein-protein interactions, and the active form of the hRad51 filament is more stimulatory than the inactive one. The current results support the hypothesis that the hRad51 conformation state has a strong effect on interaction with hRad54 and ultimately on the function of hRad54 in homologous recombination.

| S-EPMC3259846 | biostudies-other

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data