PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “Bacteriophage lambda”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 865 records · Page 48Linked to original sources

Specificity and mechanism of antitermination by Q proteins of bacteriophages lambda and 82.

Lambdoid phage late gene operons are positively regulated by genome-specific antiterminator proteins encoded by the Q gene of each phage. In this paper, we compare the activity of phage lambda and phage 82 Q proteins. Q82-mediated antitermination, like that of Q lambda, involves a transcription pause during which the regulator can modify RNA polymerase. We show that the activities of both Q82 and Q lambda are genome-specific in chasing RNA polymerase out of the early pause sites and in mediating antitermination. Finally, we show that the length of the RNA in the paused complex, or the exact position of the pause, affects the efficiency with which Q82 chases RNA polymerase out of the pause.

Bacterial Proteins↗

RNA-primed initiation sites of DNA replication in the origin region of bacteriophage lambda genome.

Using DNA molecules synthesized in the early stage of lambda phage infection, deoxynucleotides at the transition sites from primer RNA to DNA synthesis have been mapped in the 1.5 kbase area of the lambda phage genome containing the genetically defined replication origin (ori lambda). Sites in the 1-strand (the polarity of the 1-strand is 5' to 3' from the left to the right direction of the lambda phage genetic map) were distributed both inside and outside of the ori lambda, whereas the sites in the r-strand (the strand in the opposite polarity) were mainly distributed more than three hundred nucleotides apart from the ori lambda to the right. A CPuPu sequence was found at -12 to -10 region of transition sites of the r- and the 1-strands in the frequency of 80% and 70%, respectively, and over 60% of the CPuPu sequences were CAG. Properties of the transition sites are discussed in relation to the primer synthesis.

Bacteriophage lambda↗

Effect of bacteriophage lambda infection on synthesis of groE protein and other Escherichia coli proteins.

We used two-dimensional gel electrophoresis to quantitate the changes in rates of synthesis that follow phage lambda infection for 21 Escherichia coli proteins, including groE and dnaK proteins. Although total protein synthesis and the rates of synthesis of most individual E. coli proteins decreased after infection, some proteins, including groE protein, dnaK protein, and stringent starvation protein, showed increases to rates substantially above their preinfection rates. Infection by lambda Q- affected host synthesis in the same way as infection by gamma+, whereas infection by lambda N- showed no detectable effect on host synthesis. Deletion of the early genes between att and N abolished the effect, and shorter deletions in this region gave intermediate effects. By this sort of deletion mapping, we show that a large part, though not all, of the effect of lambda infection on host protein synthesis can be ascribed to the early region that contains phage genes Ea10 and ral. We compared the changes in protein synthesis after infection with the changes that occur in uninfected cells upon heat shock or amino acid starvation. The spectrum of changes that occurred on infection was very different from that seen after heat shock but quite similar to that seen during amino acid starvation. Despite this similarity of the effects of lambda infection and starvation, we did not detect any increase in the level of guanosine tetraphosphate during infection. We show that the groE protein is the same protein as B56.5 of Lemaux et al. (Cell 13:427-434, 1978) and A protein of Subramanian et al. (Eur. J. Biochem. 67:591-601, 1976).

Amino Acids↗

Fluorescence study of secondary structure of DNA within bacteriophage lambda.

Bromoacetaldehyde (BAA) was used to study the secondary structure of DNA in lambda-phage particles. It was determined that about 1% of the adenines in the intraphage lambda-DNA reacts readily with BAA, thus, they are placed in DNA sites with disturbed complementary interactions. These adenines are close to the tryptophan residues of the phage protein. Fluorescence emission of epsilon A in the intraphage DNA is dramatically quenched. This, apparently, indicates the interaction between epsilon A and Trp- and/or Tyr- and/or Met-residues of phage protein.

Bacteriophage lambda↗

[Packing of binding stained DNA strands in bacteriophage lambda].

Distribution of stainable DNA strands in phage lambda has been studied by polarized fluorescence. The effect of tight DNA-packing on fluorescence depolarization of complex dye-DNA was calculated. It is shown that stainable DNA in the phage is not concentrated in the central region. The arrangement of acridine orange molecules on the surface layers of the packed DNA is the most probable one.

Bacteriophage lambda↗

Recombinational repair of DNA damage in Escherichia coli and bacteriophage lambda.

Although homologous recombination and DNA repair phenomena in bacteria were initially extensively studied without regard to any relationship between the two, it is now appreciated that DNA repair and homologous recombination are related through DNA replication. In Escherichia coli, two-strand DNA damage, generated mostly during replication on a template DNA containing one-strand damage, is repaired by recombination with a homologous intact duplex, usually the sister chromosome. The two major types of two-strand DNA lesions are channeled into two distinct pathways of recombinational repair: daughter-strand gaps are closed by the RecF pathway, while disintegrated replication forks are reestablished by the RecBCD pathway. The phage lambda recombination system is simpler in that its major reaction is to link two double-stranded DNA ends by using overlapping homologous sequences. The remarkable progress in understanding the mechanisms of recombinational repair in E. coli over the last decade is due to the in vitro characterization of the activities of individual recombination proteins. Putting our knowledge about recombinational repair in the broader context of DNA replication will guide future experimentation.

Bacteriophage lambda↗

Enhanced activity of the bacteriophage lambda PL promoter at low temperature.

The response of the early phage lambda PL promoter to temperature was investigated. Experiments with lacZ reporter gene fusions demonstrated that the activity of the phage lambda PL promoter is inversely dependent on temperature. The bacterial DNA-binding protein integration host factor (IHF) further enhances lambda PL promoter activity at low temperature, although no apparent changes in the cellular level of IHF protein were observed at the different temperatures. IHF protein binds DNA in vitro more avidly at low temperatures. In vitro transcription assays further revealed that the temperature response of PL is the result of an intrinsic property of the promoter as well as its activation by IHF.

Bacterial Proteins↗

Transcription termination: sequence and function of the rho-independent tL3 terminator in the major leftward operon of bacteriophage lambda.

For cloning, assaying the function and sequencing terminators, we have constructed the pD12 plasmid, in which the late promotor p'R of phage lambda controls the expression of the galK gene of the pK03 plasmid of McKenney et al. (1981). The lambda tL3 terminator region was cloned in this plasmid between the promoter and the galK gene, and found to be 90-94% effective in preventing galactokinase expression in both rho+ and rho- hosts. Is is also active in vitro, both in the presence or absence of the rho factor. The termination point is located at 4320 bp to the left of the SL startpoint of the PL-RNA, just downstream of gene exo. We have sequenced 356 bp of the hitherto uncharted lambda DNA to the right of the TaqI cut, which in turn is 110 bp to the right of the b522 deletion at 63.9% lambda. The tL3 terminator has several features common to other rho-independent termination sequences, including an 81% G+C-rich region of 2X8-bp symmetry ("stem") with a 5-bp intervening "loop", partially overlapping and followed by a sequence transcribed into the pyrimidine-rich CCUUUCU-OH 3' terminus of the RNA. The termination point that follows the last U was determined by the S1 mapping technique.

Bacteriophage lambda↗

Effect of umuC mutations on targeted and untargeted ultraviolet mutagenesis in bacteriophage lambda.

Mutagenesis of phage lambda towards clear-plaque phenotype (c+----c) results in two classes of mutants that can be distinguished genetically and morphologically. Indirect mutagenesis, i.e. mutagenesis of unirradiated phage lambda c+ stimulated by the ultraviolet irradiation of the Escherichia coli host, results in mixed bursts (c/c+) of turbid wild-type and clear-plaque mutant phages. Pure bursts of lambda c mutants are induced by irradiation of the phage genome. Irradiation of both phages and host bacteria stimulates the production of the two classes of mutant clones. We show that three different mutant alleles of the E. coli umuC gene only prevent the appearance of pure bursts of clear-plaque mutants, while mixed bursts are produced at least as frequently in umuC mutants as in the umuC+ parent.

Bacteriophage lambda↗

Structure and function of mutants in the P gene of bacteriophage lambda leading to the pi phenotype.

The location of 14 independently isolated spontaneous pi A and pi B point mutants in the lambda P gene and their base exchanges were determined. It was found that the pi B mutation is one unique type mapping close to other pi A mutants. The number of possible pi A mutation sites could be estimated. The mutation sites are distributed asymmetrically in the gene. The N-terminal half of the protein is unchanged. It is assumed to be required for the interaction with the lambda O protein. The P protein can be changed by substitution of a limited number of amino acids at the C-terminus. All functional proteins of this type have pi character. pi proteins do not appear to have altered intracellular levels or stabilities as compared to wild-type P protein. The plating characteristics of our mutants on two groP- mutants located in the dnaJ and dnaK genes, respectively, are strikingly different.

Bacteriophage lambda↗

Mutations defining the operator-binding sites of bacteriophage lambda repressor.

We have characterized about 50 different amino acid substitutions in the aminoterminal domain of lambda repressor. Sixteen of these substitutions alter external side chains of the repressor and cause a substantial reduction in the affinity of the mutant repressor for operator DNA. Seven of these mutant repressors were purified and were shown to be stably folded. The strong, external repressor mutations occur near the aminoterminal end of alpha helix 2, throughout alpha helix 3, and in the aminoterminal-arm region of the repressor. These results suggest that these regions of lambda repressor are close to operator DNA in the protein-DNA complex and thus that these regions comprise the DNA-binding sites of the repressor. Our genetic results support and are completely consistent with more-detailed models of the repressor-operator interaction based on model-building (Pabo and Lewis 1982; Lewis et al. this volume) and biochemical studies (Pabo et al. 1982).

Amino Acid Sequence↗

A bacteriophage lambda vector for cloning with BamHI and Sau3A.

A phage lambda cloning vector has been constructed which contains a single site for the restriction endonuclease BamHI. Since Sau3A and Bg/II produce the same cohesive ends as BamHI, this vector can also be used to clone DNA fragments generated with either of these enzymes. We have used this vector to construct an Escherichia coli library using partial digestion with Sau3A. This vector will be most useful for applications requiring genetic analysis of cloned E. coli genes.

Bacteriophage lambda↗

Mutations in bacteriophage lambda that alter phage dependence on the htpR gene product of Escherichia coli.

Six mutants of lambda having reduced dependence on the htpR function of Escherichia coli were isolated from lambda cIts857. Burst sizes in htpRts cells at 40.5 degrees were in the range of 10 to 20 particles per cell. Mapping and complementation analysis of one of the mutants suggested that the mutation in this isolate is in gene J. Additional evidence that the mutations in most of the isolates are in J was provided by the finding that all but one of the mutants differ from the parental phage in properties pertaining to extended host range.

Bacterial Proteins↗

Use of bacteriophage lambda recombination functions to promote gene replacement in Escherichia coli.

Replacement of Escherichia coli's RecBCD function with phage lambda's Red function generates a strain whose chromosome recombines with short linear DNA fragments at a greatly elevated rate. The rate is at least 70-fold higher than that exhibited by a recBC sbcBC or recD strain. The value of the system is highlighted by gene replacement with a PCR-generated DNA fragment. The deltarecBCD::Plac-red kan replacement allele can be P1 transduced to other E. coli strains, making the hyper-Rec phenotype easily transferable.

Bacteriophage lambda↗

Involvement of DNA polymerase III in UV-induced mutagenesis of bacteriophage lambda.

It has been proposed that the mutation fixation processes stimulated by SOS induction result from an induced infidelity of DNA replication (Radman 1974). The aim of this study was to determine if mutator mutations in the E. coli DNA polymerase III might affect UV-induced mutagenesis. Using a phage lambda mutation assay which can discriminate between targeted and untargeted mutations, we show that the polC74 mutator mutation (Sevastopoulos and Glaser 1977) primarily affects untargeted mutagenesis, which occurs in a recA1 genetic background and is amplified in the recA+ genetic background. The polC74 mutation also increases the UV-induced mutagenesis of the bacterial chromosome. These results suggest that DNA polymerase III is involved in the process of UV-induced mutagenesis in E. coli.

Bacteriophage lambda↗

The pCLIP plasmids: versatile cloning vectors based on the bacteriophage lambda origin of replication.

A series of general-purpose plasmid vectors based on the phage lambda origin of replication (ori) has been constructed. Each vector consists of a backbone plasmid encoding chloramphenicol resistance (CmR) and containing a unique HaeII site into which the lacZ alpha-complementing multiple cloning site (MCS) region of an established vector was inserted. To increase the cloning potential of the inserted MCS, superfluous restriction sites in the backbone were removed by a variety of techniques. The vectors, designated pCLIP (for CmR lambda ori integration proficient) plasmids, are of medium copy number and are compatible with most other vectors in common use. A total of 17 unique restriction sites in pCLIP8, pCLIP9, pCLIP18, pCLIP19 and pCLIP23 are available for cloning. As well as possessing the usual properties of vectors, the pCLIP plasmids are able to integrate reversibly into lambda prophage by homologous recombination. Thus, cloned DNA can be maintained in single or multiple copy at will. By integrating recombinant plasmids into appropriate deletion prophages followed by induction, phage::plasmid hybrids are produced which can be manipulated as phage. These properties are demonstrated using a test recombinant plasmid, pCLIPLEU2. The pCLIP vectors are therefore useful for routine plasmid cloning, complementation analysis and applications where the ability to manipulate recombinants in plasmid, phage or prophage forms is advantageous.

Amino Acid Sequence↗

Bistability and switching in the lysis/lysogeny genetic regulatory network of bacteriophage lambda.

Bistability and switching are two important aspects of the genetic regulatory network of lambda phage. Positive and negative feedbacks are key regulatory mechanisms in this network. By the introduction of threshold values, the developmental pathway of lambda phage is divided into different stages. If the protein level reaches a threshold value, positive or negative feedback will be effective and regulate the process of development. Using this regulatory mechanism, we present a quantitative model to realize bistability and switching of lambda phage based on experimental data. This model gives descriptions of decisive mechanisms for different pathways in induction. A stochastic model is also introduced for describing statistical properties of switching in induction. A stochastic degradation rate is used to represent intrinsic noise in induction for switching the system from the lysogenic pathway to the lysis pathway. The approach in this paper represents an attempt to describe the regulatory mechanism in genetic regulatory network under the influence of intrinsic noise in the framework of continuous models.

Bacteriolysis↗

Proteolysis of bacteriophage lambda CII by Escherichia coli FtsH (HflB).

FtsH (HflB) is a conserved, highly specific, ATP-dependent protease for which a number of substrates are known. The enzyme participates in the phage lambda lysis-lysogeny decision by degrading the lambda CII transcriptional activator and by its response to inhibition by the lambda CIII gene product. In order to gain further insight into the mechanism of the enzymatic activity of FtsH (HflB), we identified the peptides generated following proteolysis of the phage lambda CII protein. It was found that FtsH (HflB) acts as an endopeptidase degrading CII into small peptides with limited amino acid specificity at the cleavage site. beta-Casein, an unstructured substrate, is also degraded by FtsH (HflB), suggesting that protein structure may play a minor role in determining the products of proteolysis. The majority of the peptides produced were 13 to 20 residues long.

ATP-Dependent Proteases↗