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The lysins of bacteriophages infecting lactic acid bacteria.

This short review highlights the complete absence of literature on lysins of bacteriophages infecting species like S. salivarius subsp. thermophilus, Pediococcus and Leuconostoc species, L. helveticus, L. acidophilus, L. plantarum and L. brevis, which are also widely used in the dairy industry. The lysins described share some similar biochemical characteristics: optimal pH and temperature, site of hydrolysis inside the peptidoglycan, and some activators and inhibitors. The cloning of the genes encoding these lysins only began in the last few years and four of them have been completely sequenced. In the future, these lysin genes could be interestingly compared to the host autolysin(s) gene(s). By contrast, the passage of phage lysins through the cytoplasmic membrane of the host cell in order to reach the peptidoglycan (via a signal sequence or the presence of a holin) seems not to be clearly resolved. The presence of a second open-reading frame upstream from the gene of the lysin, enabling a putative holin to be encoded, has already been suggested. No doubt our ever increasing knowledge about bacteriophage genome organization will help to elucidate this question. Meanwhile the obtention of a Lactococcus strain with an autolytic phenotype, using a bacteriophage lysin gene, as well as the successful use of purified PL1 lysin to obtain protoplasts of L. casei encourage us to continue to explore the field of bacteriophage lysins.

Amino Acid Sequence↗

Genetic tools for selective labeling of proteins with alpha-15N-amino acids.

A collection of genetic tools that can be used to manipulate amino acid metabolism in Escherichia coli is described. The set comprises 21 strains of bacteria, each containing a different genetic defect that is closely linked to a selectable transposon marker. These tools can be used to construct strains of E. coli with ideal genotypes for residue-specific, selective labeling of proteins with nearly any 15N-amino acid. By using strains which have been modified to contain the appropriate genetic lesions to control amino acid biosynthesis, dilution of the isotope by endogenous amino acid biosynthesis and scrambling of the label to other types of residues can be avoided.

Amino Acids↗

Dynamics of PhiX174 protein E-mediated lysis of Escherichia coli.

Expression of cloned gene E of bacteriophage PhiX174 induces lysis by formation of a transmembrane tunnel structure in the cell envelope of Escherichia coli. Ultrastructural studies of the location of the lysis tunnel indicate that it is preferentially located at the septum or at polar regions of the cell. Furthermore, the diameter and shape of individual tunnel structures vary greatly indicating that its structure is not rigid. Apparently, the contours of individual lysis tunnels are determined by enlarged meshes in the peptidoglycan net and the force produced at its orifice, by the outflow of cytoplasmic content. Once the tunnel is formed the driving force for the lysis process is the osmotic pressure difference between cytoplasm and medium. During the lysis process areas of the cytoplasmic membrane which are not tightly attached to the envelope are extended inward by the negative pressure produced during lysis. After cell lysis external medium can diffuse through the lysis tunnel filling the inner cell space of the still rigid bacterial ghosts.

Adenosine Triphosphatases↗

Evidence for a dual control of the initiation of host-cell lysis caused by phage lambda.

The timing of host-cell lysis by coliphage lambda is controlled by two separable systems. The lambdaS gene product acts as a positive effector of lysis while another protein, the lysis regulator, is an inhibitor of lysis. If the continuous function of the lysis regulator is interrupted in phage infected cells immediate lysis ensues. This lysis requires metabolic energy but not S gene activity. In contrast, S protein activity is stimulated by agents which uncouple and which block oxidative phosphorylation. The lysis regulator is thermolabile and several lines of evidence suggest that it may be the lambdarex gene product.

Bacteriolysis↗

The depression of endolysin synthesis in bacteria infected with high multiplicities of phage lambda.

The effect of multiplicity of infection was studied in Escherichia coli with lambda phage, using phage endolysin as an example of a late gene product. A very sensitive endolysin assay method was used so that the initiation time of endolysin synthesis could be more accurately determined. It was observed that high multiplicity of infection (1) increases the rate of lysogenization, (2) progressively delays lysis time, and (3) significantly delays and reduces the synthesis of endolysin in lamdacIII+ cII+ -infected cells. The extent of delay and reduction in endolysin synthesis increases with increasing multiplicity. In contrast, lamdacIII67cII68-infected cells show no delay in endolysin synthesis at high multiplicity of infection when compared with the lamdacIII+ cII+ -infected cells. The results suggest that (1) the expression of cIII and cII genes is multiplicity dependent, (2) high multiplicity of infection enhances the expression of the cIII and cII genes, and (3) the expression of the cIII and cII genes interferes with the expression of the late genes. A model to explain how the expression of the cIII and cII genes interferes with the expression of the late genes is proposed.

Bacteriolysis↗

Envelope mutation promoting autolysis in Salmonella typhimurium.

Two strains independently isolated in Salmonella typhimurium display abnormal autolytic activity when nutrient broth becomes alkaline. They also show increased sensitivity to deoxycholate, EDTA, and sodium dodecyl sulfate. Response to acridine orange remains normal. In both strains a single stable mutation is responsible for all the changes. The same gene, called envD, appears to be involved in both mutant strains. envD has been located at minute 33 of the Salmonella genetic map, between markers sucA and nadA, very close to the latter. envD also affects morphological characteristics of the cells. Many mutant cells are shorter than wild type bacteria, and appear frequently associated in short chains of 4 to 10 cells. Furthermore, envD mutants display division by septation under conditions that preclude its observation in wild type strains.

Bacteriolysis↗

Colicin M is only bactericidal when provided from outside the cell.

The colicin M structural gene, cma, was subcloned in a vector which allowed temperature-inducible control of its expression. Induction of expression of cma in colicin M uptake proficient strains was lethal for the host cell when the colicin M immunity protein was not present. In liquid culture cells lysed, and no colonies were formed on solid media. These effects were not observed in mutants defective in the colicin receptor (FhuA) or uptake functions (TonB, TolM), nor in wild-type cells treated with trypsin prior to induction of cma expression. It was concluded that cytoplasmic colicin M is not toxic for the producing cell. To exert a lethal effect the colicin has to enter the cell from outside. Cells expressing cma released small amounts of colicin M.

Bacterial Proteins↗

Effect of aminoglycoside antibiotics on the autolytic enzyme of Streptomyces griseus.

The isolated cell wall of Streptomyces griseus 52-1 strain labelled with fluorescein isothiocyanate (FITC) and containing wall-bound autolytic enzyme was lysed as a function of different cations. The autolysis was accelerated by aminoglycoside antibiotics (streptomycin and the structurally closely related neomycin) which have a polycationic character. Since this strain is a streptomycin producer it is suggested that streptomycin may have a regulatory function on autolysis.

Amidohydrolases↗

Synchronous division induced in Escherichia coli K12 by gemts mutants of phage Mu.

Infection with the bacteriophage mutant Mu c+ gemts2 at 42 degrees C induces synchrony in cell division in cultures of Escherichia coli K12. This synchrony may last for several cycles and is not only due to selection since synchronization is observed even when bacterial survival to the infection is over 80% as in lysogens for Mu c+ gemts2. The mechanism by which synchrony is induced is not known, but since the product of Mu gene gem (previously called lig) has been shown to interact with the enzymatic system in the bacteria controlling the degree of DNA supercoiling, the phenomenon could be a consequence of this interaction.

Ampicillin Resistance↗

Translational regulation of the lysis gene in RNA bacteriophage fr requires a UUG initiation codon.

Single nucleotide substitutions identify a UUG triplet as the initiation codon of the lysis gene in RNA bacteriophage fr. This initiation codon is non-functional in de novo initiation but is activated by translational termination at the overlapping coat gene. The UUG initiation codon is crucial for gene regulation in the phage, as it excludes uncontrolled access of ribosomes to the start of the lysis gene. Replacement of UUG by either GUG or AUG results in the loss of genetic control of the lysis gene. A model is presented in which initiation factor IF3 proofreads de novo initiation at UUG codons.

Bacteriolysis↗