Cell membrane integrity and lysis in Lactococcus lactis: the detection of a population of permeable cells in post-logarithmic phase cultures

Niven, Gordon W.; Mulholland, Francis

doi:10.1046/j.1365-2672.1997.00316.x

Cited by 50 publications

(50 citation statements)

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“…Phenotypes of the resulting MG1363yjgB, MG1363acmByjgB, and MG1363acmAacmByjgB mutants (Table 1) were analyzed by a number of custom assays used for characterization of pepti- (17). These included analysis of cellular growth rate, measurement of cell autolysis in the stationary phase both by decrease in optical density and by activity of concomitantly released intracellular peptidase PepX, and analysis of membrane permeabilization in the stationary phase with the fluorescent DNA marker propidium iodide (25). The assays demonstrated no visible phenotypic change associated with yjgB inactivation in either of the created mutants compared to the wild type (data not shown).…”

Section: Resultsmentioning

confidence: 99%

Lactococcus lactis Gene yjgB Encodes a γ- d -Glutaminyl- l -Lysyl- Endopeptidase Which Hydrolyzes Peptidoglycan

Redko

Courtin

Mézange

et al. 2007

Appl Environ Microbiol

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YjgB is one of five peptidoglycan hydrolases previously identified in Lactococcus lactis. Analysis of its amino acid sequence revealed that YjgB contains an NlpC/P60 domain, whereas no specific cell wall binding domain or motif could be identified. The NlpC/P60 family is characterized by three conserved residues, a cysteine, a histidine, and a polar residue. In agreement with the presence of a Cys residue in the catalytic site of YjgB, its enzymatic activity was enhanced in the presence of dithiothreitol. Peptidoglycan-hydrolyzing activity of YjgB was detected in growing cells of an L. lactis strain overexpressing YjgB, as revealed by the presence of disaccharide (DS)-dipeptide in the muropeptide composition of the overexpressing strain. YjgB hydrolyzes the peptide chains of L. lactis muropeptides between ␥-D-Gln and L-Lys residues. Its hydrolytic activity was detected on DSs with tetra-and pentapeptide chains, whereas hydrolytic activity was very low on DS-tripeptides. Thus, we demonstrated that YjgB is an endopeptidase which cleaves ␥-D-Gln-L-Lys bonds in peptide chains of L. lactis peptidoglycan.The cell integrity of gram-positive bacteria is ensured by a cell wall which consists mainly of peptidoglycan, an aminosugar polymer cross-linked with peptide bridges. Peptidoglycan metabolism is a necessary process in cell wall remodeling during growth and cell division, during which peptidoglycan hydrolysis is balanced with the synthesis of new polymer blocks (9, 31). While cell integrity is in this case maintained, it is compromised during cellular autolysis. Both processes require the action of enzymes called peptidoglycan hydrolases (PGHs), which catalyze cleavage of peptidoglycan sugar or peptide chains. In lactic acid bacteria such as Lactococcus lactis, which is used as a dairy starter, PGH activity during cheese ripening leads to cellular autolysis and release of the cellular content to the cheese curd. This provides the pool of enzymes acting in the development of organoleptic properties of cheese (22). Therefore, control of PGH activity in L. lactis bears the potential for control of cheese ripening in industrial conditions. The PGH complement of L. lactis was analyzed previously and comprises five PGHs (16). It includes AcmA, AcmB, and AcmC, all of which exhibit N-acetylglucosaminidase hydrolytic specificity (16,17,32). AcmA is the major autolysin of L. lactis; it is involved in cell separation after cell division and in cellular autolysis when cells reach the stationary phase (6). The Nterminal part of AcmA contains the catalytic domain, while the C-terminal part comprises three LysM domains involved in cell wall binding (33). AcmB, the second PGH of L. lactis, is built of three domains: an S/T/P/N-rich domain at its N terminus followed by the catalytic domain and a C-terminal part of unknown function. AcmB contributes to cellular autolysis to a lesser extent than AcmA and does not participate in cell separation (17). The third L. lactis PGH, AcmC, does not contain any specific cell wall binding domain an...

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Section: Resultsmentioning

confidence: 99%

Lactococcus lactis Gene yjgB Encodes a γ- d -Glutaminyl- l -Lysyl- Endopeptidase Which Hydrolyzes Peptidoglycan

Redko

Courtin

Mézange

et al. 2007

Appl Environ Microbiol

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“…Furthermore, the membrane permeability of L. lactis was measured with PI and carboxyfluorescein diacetate (cFDA) using spectrofluorimetry and fluorescence microscopy in a study of viability after various stress treatments (6). Also, permeable fractions in post-logarithmic-phase cultures of L. lactis were measured with PI using spectrofluorimetry (22). Finally, permeabilization by bile salts and acid with concomitant cell death of various lactic acid bacteria was measured with PI, TOTO-1, and cFDA (5).…”

Section: Discussionmentioning

confidence: 99%

“…Alternative methodologies for extraction such as using a cheese press and using hypertonic buffers have been tried (30). However, to date no really good method is available, and an understanding of the role of lysis in cheese ripening is still lacking (8,22).…”

mentioning

confidence: 99%

Fluorescent Method for Monitoring Cheese Starter Permeabilization and Lysis

Bunthof

Schalkwijk

Meijer

et al. 2001

Appl Environ Microbiol

106

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“…Effect on Bacterial Membrane Integrity and Growth. The effect of 200 nM amine-and carboxyl-PEG-QDs on cell membrane integrity was probed by FCM using PI, a fluorescent nucleic acid stain that does not penetrate cells with intact membranes, while staining cells with damaged or leaky membranes (34). FCM analysis of PI treated cells demonstrated that both amine-and carboxyl-PEG-QDs induce an increase in the number of damaged/leaky membranes of C. metallidurans cells (Figure 3).…”

Section: Mops)mentioning

confidence: 99%

Amine- and Carboxyl- Quantum Dots Affect Membrane Integrity of Bacterium Cupriavidus metallidurans CH34

Startchev

Roberts

2009

Environ. Sci. Technol.

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The present study examines the interaction of amine-and carboxyl-PEG core/shell quantum dots (QDs) with metal resistant bacterium Cupriavidus metallidurans CH34. The evolution of the number of QDs, their hydrodynamic radius, diffusion coefficients, and single particle fluorescence were characterized before and during the contact with bacterium by fluorescence correlation spectroscopy (FCS). The obtained results showed that at nanomolar concentrations the amine-and carboxyl-PEG-QDs with average hydrodynamic radiuses of 16.4 and 13.5 nm, form stable dispersions in the absence and presence of 15 mgC L -1 HA. The decrease of the number of fluorescent particles in the bacterial medium, determined by FCS, together with the increase of the fluorescence of bacterial cells over the background, found by flow cytometry (FCM), demonstrated the association of QDs to C. metallidurans. Furthermore, QDs enhanced the level of the reactive oxygen species in the bacterial cells and augmented the percentage of the cells with damaged and leaky membranes as probed by FCM in combination with 5-(and-6)-carboxy-2′7′-dichlorodihydrofluorescein diacetate and propidium iodide stains. No difference in the behavior of amine-and carboxyl-PEG-QDs was found, suggesting that different functional groups in the surface coating have no effect on bacterium-QD interactions under the studied conditions. The presence of HA does not affect the hydrodynamic characteristics of the functionalized QDs, but prevented the damage to the bacterial membrane. The slight decrease in the bacterial growth found after exposure of C. metallidurans to these QDs was attributed to the nanoparticles themselves rather the cadmium, zinc, or selenium ions released from the QDs.

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Cell membrane integrity and lysis in Lactococcus lactis: the detection of a population of permeable cells in post-logarithmic phase cultures

Cited by 50 publications

References 3 publications

Lactococcus lactis Gene yjgB Encodes a γ- d -Glutaminyl- l -Lysyl- Endopeptidase Which Hydrolyzes Peptidoglycan

Lactococcus lactis Gene yjgB Encodes a γ- d -Glutaminyl- l -Lysyl- Endopeptidase Which Hydrolyzes Peptidoglycan

Fluorescent Method for Monitoring Cheese Starter Permeabilization and Lysis

Amine- and Carboxyl- Quantum Dots Affect Membrane Integrity of Bacterium Cupriavidus metallidurans CH34

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