Mechanism of autolysis of Neisseria gonorrhoeae

Hebeler, B H; Young, Frank E.

doi:10.1128/jb.126.3.1186-1193.1976

Cited by 38 publications

(24 citation statements)

References 13 publications

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“…If these relatively few glycosidic bonds are lysozyme insensitive yet susceptible to autolysins, as has been observed with Lactobacillus fermentum (14), then the pH dependence of the lysis reaction reflects the activity of the autolytic enzymes of S. mutans GS5 and not of the lysozyme. Similar pH-dependent effects have been observed for the autolysins of other bacterial species (7,10,23). Autolytic as well as lysozyme activity has been reported to vary with the growth phase (17), and in this study peaks of lytic activity were noted during exponential growth of the organisms (Fig.…”

Section: Discussionsupporting

confidence: 85%

Bacteriolysis of Streptococcus mutans GS5 by lysozyme, proteases, and sodium thiocyanate

Wilkens¹,

Goodman²,

Mackay³

et al. 1982

Infect Immun

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Streptococcus mutans GS5 was grown in a synthetic medium containing radioactive thymidine to monitor cell lysis by assay of the release of DNA. Bacteriolysis was achieved by sequential treatment of the cells with either hen egg white lysozyme and sodium thiocyanate or a combination of hen egg white lysozyme and a proteolytic enzyme followed by addition of the thiocyanate. In the absence of sodium thiocyanate, a small percentage of the total macromolecular thymidine was released in control reaction mixtures during incubation. This amount of released DNA more than doubled in trypsin-treated cells, but the inclusion of lysozyme in reaction mixtures prevented assay of the DNA. Lysis was found to be optimal in the late log phase of growth and was dependent on the concentrations of both lysozyme and protease. Concentrations of trypsin or chymotrypsin as low as 0.01 Fxg/ml were found to be effective in enhancing the lytic process. The addition of protease to lysozyme-inorganic salt reaction mixtures altered both the pH and ionic strength profiles of cell lysis. At pHs of 5.5 or lower, both the lysozyme-NaSCN and the lysozyme-trypsin-NaSCN systems were inactive in mediating lysis. The loss of insoluble cell wall peptidoglycan by lysozyme treatment was pH independent and did not appear to be affected by the addition of protease. Either diluted whole saliva or neutrophil extracts could replace trypsin to enhance cell lysis further.Detailed studies of the bacteriolysis of a variety of microorganisms exhibiting different degrees of susceptibility to lysozyme and inorganic salts are required for the ultimate understanding of the mechanism of cell lysis. In previous communications from our laboratories, we have examined Streptococcus mutans BHT, a serotype b strain (20), which is very sensitive to the lytic action of hen egg white lysozyme (EC 3.2.1.17) (HEWL) and the sodium salts of thiocyanate, bicarbonate, chloride, and fluoride (9,21,22). In this study, we report on the bacteriolysis of S. mutans GS5, a serotype c strain, which is less susceptible to lysis than strain BHT is. During investigation of the lysis of strain GS5 and other oral microorganisms, it has become apparent that a range of lytic sensitivities does indeed exist. Moreover, in this communication, we report on the enhancement of lysozymeinorganic salt lysis by proteases. Preliminary data are also given which demonstrate that the proteases can be replaced in the assay mixtures by either diluted whole saliva or diluted neutrophil extracts, perhaps suggesting a physiological role for the lysozyme-protease-salt antibacterial lytic system in the oral cavity. MATERIALS AND METHODS Biochemicals. HEPES [N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid], MES [2-(N-morpholino)ethanesulfonic acid], PIPES [piperazine-N,N'bis(2-ethanesulfonic acid)], and CHES [2-(N-cyclohexylamino)ethanesulfonic acid] were obtained from Calbiochem, La Jolla, Calif. Tris, sodium thiocyanate, magnesium sulfate, and trichloroacetic acid were purchased from Fisher Scientific Co., Pittsbur...

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Section: Discussionsupporting

confidence: 85%

Bacteriolysis of Streptococcus mutans GS5 by lysozyme, proteases, and sodium thiocyanate

Wilkens¹,

Goodman²,

Mackay³

et al. 1982

Infect Immun

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“…The tendency of Ngo to autolyse has long been observed and attributed to murein hydrolases (Hebeler and Young, 1976a;Rosenthal, 1979). Specifically, N-acetylmuramyl-i-alanine amidase was proposed to be a major autolysin of Ngo (Hebeler and Young, 1976b;Chapman and Perkins, 1983). The controlled autolysis of a part of the population may be seen as a general mechanism by which the gonococcus can set DNA free, thus making its gene pool available to competent neighbouring Ngo able to take the DNA up in a species-specific manner.…”

Section: Discussionmentioning

confidence: 99%

Tetrapac (tpc), a novel genotype of Neisseria gonorrhoeae affecting epithelial cell invasion, natural transformation competence and cell separation

Fussenegger

Kahrs

Facius

et al. 1996

Molecular Microbiology

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We characterized a novel mutant phenotype (tetrapac, tpc) of Neisseria gonorrhoeae (Ngo) associated with a distinctive rough-colony morphology and bacterial growth in clusters of four. This phenotype, suggesting a defect in cell division, was isolated from a mutant library of Ngo MS11 generated with the phoA minitransposon TnMax4. The tpc mutant shows a 30% reduction in the overall murein hydrolase activity using Escherichia coli murein as substrate. Tetrapacs can be resolved by co-cultivation with wild-type Ngo, indicating that Tpc is a diffusible protein. Interestingly, Tpc is absolutely required for the natural transformation competence of piliated Ngo. Mutants in tpc grow normally, but show a approximately 10-fold reduction in their ability to invade human epithelial cells. The tpc sequence reveals an open reading frame of approximately 1 kb encoding a protein (Tpc) of 37 kDa. The primary gene product exhibits an N-terminal leader sequence typical of lipoproteins, but palmitoylation of Tpc could not be demonstrated. The ribosomal binding site of tpc is immediately downstream of the translational stop codon of the folC gene coding for an enzyme involved in folic acid biosynthesis and one-carbon metabolism. The tpc gene is probably co-transcribed from the folC promoter and a promoter located within the folC gene. The latter promoter sequence shares significant homology with E. coli gearbox consensus promoters. All three mutant phenotypes, i.e. the cell separation defect, the transformation deficiency and the defect in cell invasion can be restored by complementation of the mutant with an intact tpc gene. To some extent the tcp phenotype is reminiscent of iap in Listeria, lytA in Streptococcus pneumoniae and lyt in Bacillus subtilis, all of which are considered to represent murein hydrolase defects.

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“…Gonococci are notoriously fragile and do not survive under several common laboratory conditions, including growth in the stationary phase (Morse and Bartenstein, 1974;Elmros et al, 1976a). As gonococci have been shown to express at least three autolytic activities (Hebeler and Young, 1976;Sinha and Rosenthal, 1980;Chapman and Perkins, 1983), the rapid decrease in viability in stationary phase is thought to be due to autolysis. The timing of the fall can be affected by the amount of carbon source supplied, suggesting that the cells begin to lyse when nutrients are exhausted (Morse and Bartenstein, 1974).…”

Section: Phenotypic Characterization Of An Atla Mutantmentioning

confidence: 99%

A peptidoglycan hydrolase similar to bacteriophage endolysins acts as an autolysin in Neisseria gonorrhoeae

Dillard

Seifert

1997

Molecular Microbiology

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SummaryWe have identified a gene encoding an autolysin (atlA) from Neisseria gonorrhoeae. The deduced amino acid sequence of AtlA shows significant similarity to the peptidoglycan degrading transglycosylases (endolysins) of bacteriophages lambda and P2, suggesting that the encoded protein also functions in peptidoglycan hydrolysis. An atlA mutant was identical to the wild-type strain in exponential growth rate, but demonstrated reduced lysis and peptidoglycan turnover in the stationary phase of growth. When transferred into a buffer solution, at a pH non-permissive for other gonococcal autolysins, an autolytic activity was detectable in the wild-type strain that was not present in the mutant. The most dramatic phenotype of the mutant occurred after extended time in stationary phase. After approximately 16 h in stationary phase, both strains underwent an apparent replication event, after which the wild-type strain died rapidly whereas the atlA mutant survived considerably longer. Even after both the wild-type and mutant cells were dead, many of the mutant cells maintained intact morphology, whereas the wild-type cells were lysed. These results suggest that AtlA is a peptidoglycan transglycosylase related to bacteriophage endolysins and acts as an autolysin in the stationary phase.

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Mechanism of autolysis of Neisseria gonorrhoeae

Cited by 38 publications

References 13 publications

Bacteriolysis of Streptococcus mutans GS5 by lysozyme, proteases, and sodium thiocyanate

Bacteriolysis of Streptococcus mutans GS5 by lysozyme, proteases, and sodium thiocyanate

Tetrapac (tpc), a novel genotype of Neisseria gonorrhoeae affecting epithelial cell invasion, natural transformation competence and cell separation

A peptidoglycan hydrolase similar to bacteriophage endolysins acts as an autolysin in Neisseria gonorrhoeae

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