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At least 1,063 records · Page 59Linked to original sources
PubMed1998

Helicobacter pylori containing only cytoplasmic urease is susceptible to acid.

Helicobacter pylori, an important etiologic agent in a variety of gastroduodenal diseases, produces large amounts of urease as an essential colonization factor. We have demonstrated previously that urease is located within the cytoplasm and on the surface of H. pylori both in vivo and in stationary-phase culture. The purpose of the present study was to assess the relative contributions of cytoplasmic and surface-localized urease to the ability of H. pylori to survive exposure to acid in the presence of urea. Toward this end, we compared the acid resistance in vitro of H. pylori cells which possessed only cytoplasmic urease to that of bacteria which possessed both cytoplasmic and surface-localized or extracellular urease. Bacteria with only cytoplasmic urease activity were generated by using freshly subcultured bacteria or by treating repeatedly subcultured H. pylori with flurofamide (1 microM), a potent, but poorly diffusible urease inhibitor. H. pylori with cytoplasmic and surface-located urease activity survived in an acid environment when 5 mM urea was present. In contrast, H. pylori with only cytoplasmic urease shows significantly reduced survival when exposed to acid in the presence of 5 mM urea. Similarly, Escherichia coli SE5000 expressing H. pylori urease and the Ni2+ transport protein NixA, which expresses cytoplasmic urease activity at levels similar to those in wild-type H. pylori, survived minimally when exposed to acid in the presence of 5 to 50 mM urea. We conclude that cytoplasmic urease activity alone is not sufficient (although cytoplasmic urease activity is likely to be necessary) to allow survival of H. pylori in acid; the activity of surface-localized urease is essential for resistance of H. pylori to acid under the assay conditions used. Therefore, the mechanism whereby urease becomes associated with the surface of H. pylori, which involves release of the enzyme from bacteria due to autolysis followed by adsorption of the enzyme to the surface of intact bacteria ("altruistic autolysis"), is essential for survival of H. pylori in an acid environment. The ability of H. pylori to survive exposure to low pH is likely to depend on a combination of both cytoplasmic and surface-associated urease activities.

Benzamides↗
PubMed2006

The atlA operon of Streptococcus mutans: role in autolysin maturation and cell surface biogenesis.

The Smu0630 protein (AtlA) was recently shown to be involved in cell separation, biofilm formation, and autolysis. Here, transcriptional studies revealed that atlA is part of a multigene operon under the control of at least three promoters. The morphology and biofilm-forming capacity of a nonpolar altA mutant could be restored to that of the wild-type strain by adding purified AtlA protein to the medium. A series of truncated derivatives of AtlA revealed that full activity required the C terminus and repeat regions. AtlA was cell associated and readily extractable from with sodium dodecyl sulfate. Of particular interest, the surface protein profile of AtlA-deficient strains was dramatically altered compared to the wild-type strain, as was the nature of the association of the multifunctional adhesin P1 with the cell wall. In addition, AtlA-deficient strains failed to develop competence as effectively as the parental strain. Mutation of thmA, which can be cotranscribed with atlA and encodes a putative pore-forming protein, resulted in a phenotype very similar to that of the AtlA-deficient strain. ThmA was also shown to be required for efficient processing of AtlA to its mature form, and treatment of the thmA mutant strain with full-length AtlA protein did not restore normal cell separation and biofilm formation. The effects of mutating other genes in the operon on cell division, biofilm formation, or AtlA biogenesis were not as profound. This study reveals that AtlA is a surface-associated protein that plays a critical role in the network connecting cell surface biogenesis, biofilm formation, genetic competence, and autolysis.

Bacterial Proteins↗
PubMed1970

Formation of bacteriolytic enzymes in batch and continuous culture of Staphylococcus aureus.

The formation of bacteriolytic enzymes of Staphylococcus aureus, with special reference to strain M18, was investigated under a variety of conditions. The bacteriolytic activity was tested by using whole cells of Micrococcus lysodeikticus as a substrate. Complex media were required for production, and a Casein Hydrolysate-Yeast Extract medium (CCY(I)) was superior to Brain Heart Infusion and Trypticase Soy Broth. The optimal pH level for production was 7.0. Effective oxygenation and exchange of the beta-glycerophosphate of the CCY(I) medium for glucose increased the rates of growth and autolysis and the rate of appearance of extracellular bacteriolytic enzymes. However, the extracellular lytic activity decreased more rapidly at the end of the growth period than under the standard culture conditions. The appearance of inhibitor(s), probably derived from autolysis, might be responsible for this rapid decrease. The highest yields were obtained in a continuous process in which the activity was almost twice that of batch cultures grown under the same conditions. The bacteriolytic activity produced in continuous culture had a considerably increased stability in the purification process. The advantage of producing unstable bacterial proteins in continuous culture under controlled growth conditions is discussed.

Bacteriolysis↗
PubMed1973

Some properties of the autolytic N-acetylmuramidase of Lactobacillus acidophilus.

The autolytic N-acetylmuramidase present in Lactobacillus acidophilus strain 63 AM Gasser has an optimal pH between 5 and 6 when lysing intact cells or isolated cell walls. Cellular lysis at pH 5 is two to four times more rapid in citrate buffer of 0.01 M and 0.5 M or higher than in 0.1 M acetate buffer. It seems that sulfhydryl groups are required for both cell and wall autolysis. Heavy metal ions and p-chloro-mercuribenzoate, at low concentrations, are powerful inhibitors. Ethylenediaminetetraacetic acid stimulates cellular but not wall autolysis in acetate buffer to the level obtained in citrate buffer. The possible involvement of sulfhydryl groups in a mechanism of control of cellular autolytic activity is discussed. The autolytic enzyme, although unstable in solution at 37 C, can be extracted from walls by the use of solutions of bovine serum albumin (100 mug/ml) in 0.01 N NaOH. Soluble enzyme extracted from walls rebinds on to sodium decylsulfate-treated walls, but three times as much of the wall material is required to completely re-adsorb the activity.

Acetates↗
PubMed1975

Altered crystal violet permeability and lytic behavior in antibiotic-resistant and -sensitive mutants of Neisseria gonorrhoeae.

Wild-type, antibiotic-resistant and hypersensitive isogenic strains of Neisseria gonorrhoeae were studied for uptake of crystal violet, rates of autolysis, and response to lysozyme. Total uptake of crystal violet was similar in all strains at 0 C but varied significantly at 37 C. Mutation at the nonspecific resistance locus ery resulted in relative impermeability to crystal violet at 37 C, as compared to wild type. The penetration barrier to crystal violet at 37 C was overcome by addition of 5 mM ethylenediaminetetraacetic acid. Mutation at ery also resulted in reduced rates of autolysis and reduced sensitivity to high concentrations of lysozyme under conditions of divalent cation (Mg2+) depletion. In contrast, mutation at the nonspecific drug hypersensitivity locus env resulted in increased uptake of crystal violet at 37 C, due to increased binding of dye to crude envelope as well as increased penetration into cytoplasm. The env mutants were also more rapidly autolytic and more sensitive to lysozyme than wild type in the absence of Mg2+. These results suggest that the cell envelopes of ery mutants are more stable and less permeable and those of env mutants are less stable and more permeable than wild-type strains.

Anti-Bacterial Agents↗
PubMed1978

Autolytic defective mutant of Streptococcus faecalis.

Properties of a variant of Streptococcus faecalis ATCC 9790 with defective cellular autolysis are described. The mutant strain was selected as a survivor from a mutagenized cell population simultaneously challenged with two antibiotics which inhibit cell wall biosynthesis, penicillin G and cycloserine. Compared to the parental strain, the mutant strain exhibited: (i) a thermosensitive pattern of cellular autolysis; (ii) an autolytic enzyme activity that had only a slightly increased thermolability when tested in solution in the absence of wall substrate; and (iii) an isolated autolysin that had hydrolytic activity on isolated S. faecalis wall substrate indistinguishable from that of the parental strain, but that was inactive when tested on walls of Micrococcus lysodeikticus as a substrate. These data indicate an alteration in the substrate specificity of the autolytic enzyme of the mutant which appears to result from the synthesis of an altered form of autolytic enzyme.

Bacteriolysis↗
PubMed1978

Regulation of bacterial cell walls: correlation between autolytic activity and cell wall turnover in Staphylococcus aureus.

Cell wall turnover was examined in parent and mutant strains of Staphylococcus aureus. Peptidoglycan and teichoic acid were observed to undergo turnover in the wild-type strain during exponential growth; however, the rate of turnover did not decrease when the growth rate slowed, as the culture entered stationary phase. Isolated native cell walls and crude soluble autolytic enzyme were prepared from cells harvested during exponential and postexponential phases of growth. Native cell walls from both phases of growth autolyzed in buffer at identical rates; similarily, crude soluble enzyme from both preparations degraded radioactive cell walls at the same rate. Therefore, the activity of the autolysin in both exponential and postexponential cells was similar. The autolysis of whole cells of a mutant tar-1 was enhanced by 1.0 M NaCl. When 1.0 M NaCl was present under growing conditions, the rate of cell wall turnover was greatly increased. The presence of chloramphenicol, which inhibits whole-cell autolysis, also inhibited turnover. Analysis of the cell wall material recovered from spent medium revealed products consistent with the known mode of action of the endogenous autolysin. It is concluded that cell wall turnover in S. aureus is independent of the stage of culture growth but is dependent instead on the activity of the autolysin.

Bacteriolysis↗
PubMed1978

Cell wall composition and associated properties of methicillin-resistant Staphylococcus aureus strains.

Methicillin-resistant (MR) Staphylococcus aureus strains have previously been reported to be deficient in surface negative charge; this has been correlated with methicillin resistance and ascribed to a deficiency of teichoic acid at the cell surface (A. W. Hill and A. M. James, Microbios 6:157-167, 1972). Teichoic acid was present in walls of MR organisms as revealed by appreciable phosphate levels and detection of ribitol residues. Phosphate levels in walls from five MR strains (0.54 to 0.77 mumol/mg of wall) were lower than in three unrelated methicillin-sensitive (MS) strains (0.86 to 1.0 mumol/mg of wall). However, two MS strains derived from two of the MR strains had wall phosphate levels very similar to those of the MR strains. No evidence for unusual wall polymers was found. Simple deficiency of wall teichoic acid does not result in methicillin resistance since an independently isolated teichoic acid-deficient strain (0.1 mumol of phosphate per mg of wall) was not methicillin resistant. In studies of biological properties possibly related to wall teichoic acid, it was discovered that walls isolated from MR organisms grown in the presence of methicillin autolyzed more rapidly than those isolated from organisms grown in the absence of the drug. Since methicillin resistance is enhanced by NaCl and suppressed by ethylenediaminetetraacetate, the effects of these compounds on autolysis of isolated walls were studied. NaCl (1.0 M) and ethylenediaminetetraacetate (1.0 mM) inhibited the autolysis of walls isolated from MR and MS strains. An MR strain bound phage 47, 52A, and 3A only slightly less well than their respective propagating strains.

Amino Acids↗
PubMed1981

Effects of furazlocillin, a beta-lactam antibiotic which binds selectively to penicillin-binding protein 3, on Escherichia coli mutants deficient in other penicillin-binding proteins.

Furazlocillin binds selectively to penicillin-binding protein 3 (PBP-3), prevents septation of Escherichia coli, and allows the cells to form long filaments without lysis. The effect of furazlocillin on the morphology, autolysis, and murein synthesis of E. coli mutants deficient in either PBP-1A, PBP-1Bs, or PBP-2 was studied. The results reveal that PBP-1A and PBP-1Bs functions are not equivalent since furazlocillin affects the morphology, autolysis, and murein synthesis of PBP1A- mutants quite differently from that of PBP-1Bs mutants. Different "PBP-2-" mutants were found to respond to furazlocillin in dramatically different ways: strain LS-1 cells formed elongated rods with a central bulge which eventually lysed, whereas SP6 cells formed stable "barbells" in which the two daughter cells were well separated but remained connected by a thick central region.

Azlocillin↗
PubMed1985

Involvement of autolysin in cellular lysis of Bacillus subtilis induced by short- and medium-chain fatty acids.

The addition of saturated C6, C8, C10, and C12 fatty acids appeared to lyse actively growing cells of Bacillus subtilis 168, as judged by a decrease in the optical density of the culture. Of these fatty acids, dodecanoic acid was the most effective, with 50% lysis occurring in about 30 min at a concentration of 0.5 mM. These conditions also decreased the amount of peptidoglycan estimated by the incorporated radioactivity of N-acetyl-D-[1-14C]glucosamine. At concentrations above 1 mM, however, bacterial lysis was not extensive. Dodecanoic acid did not affect autolysis of the cell wall. The lytic action of dodecanoic acid was greatly diminished in cells in which protein synthesis was inhibited and in an autolytic enzyme-deficient mutant. The results suggest that fatty acid-induced lysis of B. subtilis 168 is due to the induction of autolysis by an autolytic enzyme rather than massive solubilization of the cell membrane by the detergent-like action of the fatty acids.

Amidohydrolases↗
PubMed1986

Role of cell cohesion in Myxococcus xanthus fruiting body formation.

Dsp mutants of Myxococcus xanthus have a complex phenotype with abnormal cell cohesion, social motility, and development. All three defects are the result of a single mutation in the dsp locus, a region of DNA about 14 kilobases long. Cohesion appears to play a central role in social motility, since nonsocial mutants exhibit weak agglutination or, in the case of Dsp cells, no agglutination (L. J. Shimkets, J. Bacteriol. 166:837-841, 1986). However, Dsp cells can be agglutinated by cohesive strains of M. xanthus. This provided the opportunity to examine the role of cohesion during development by comparing the developmental phenotype of Dsp cells with that of Dsp cells mixed with cohesive strains. Dsp mutants were unable to complete any of the developmental behaviors: aggregation, fruiting body formation, developmental autolysis, and sporulation. Contact with cohesive strains seemed to restore some developmental characteristics to the Dsp cells. When allowed to develop with wild-type cells, Dsp cells accumulated in fruiting bodies and underwent developmental autolysis, but did not form a significant portion of the spore population. Igl mutants, which may be similar to the previously described frizzy mutants, are cohesive strains that are unable to form fruiting bodies. Mixing Igl cells with Dsp cells under developmental conditions resulted in fruiting body formation, although the Dsp cells were unable to form significant levels of myxospores. In spite of their inability to sporulate under developmental conditions, Dsp mutants did not appear to be defective in the sporulation process. In fact, they formed normal levels of myxospores in response to the chemical inducer glycerol.

Agglutination↗
PubMed1988

Insertional inactivation of the major autolysin gene of Streptococcus pneumoniae.

The lytA gene encoding the major pneumococcal autolysin (N-acetylmuramoyl-L-alanine amidase) was inactivated by inserting the 2-kilobase MspI fragment of pE194 containing the staphylococcal ermC gene. Stable autolysis-deficient (Lyt-) mutants and their isogenic Lyt+ parents were used in experiments designed to test possible physiological functions of the amidase. No autolysis could be induced in the mutants grown at 37 degrees C by deoxycholate, by incubation in stationary phase, or by treatment with penicillin. On the other hand, the Lyt- mutants exhibited normal growth rates and yields and normal adaptive responses during shifts from one growth temperature or nutritional condition to another. There was no evidence for impeded cell separation (chain formation). Colonies of Lyt- insertional mutants produced normal hemolytic zones on blood agar; they showed normal (high) levels of competence for genetic transformation. Lyt- mutants were also able to produce type 3 and 6 capsular polysaccharides, and such strains showed the same degree of virulence in mice as did the isogenic Lyt+ parent. The physiological function(s) of the amidase remains a puzzle.

Amidohydrolases↗
PubMed1991

Complementation of the lytD1 mutation of Escherichia coli by either the cI or cro gene of bacteriophage lambda.

The lytD1 mutant of Escherichia coli exhibits temperature-sensitive growth which is attributed to cellular autolysis at the restrictive temperature. Either of two cloned phage lambda genes, identified as cI and cro, suppressed the lytD1(Ts) lysis phenotype, suggesting that lytD encodes a DNA-binding protein with a DNA-binding specificity similar to that of CI and Cro. LytD may be a repressor of a gene(s) involved in cellular autolysis.

Bacteriophage lambda↗
PubMed1993

A cdc-like autolytic Saccharomyces cerevisiae mutant altered in budding site selection is complemented by SPO12, a sporulation gene.

LYT1 is an essential gene for the growth and morphogenesis of Saccharomyces cerevisiae. A detailed characterization of mutants carrying the lyt1-1 allele showed that this mutation was recessive and pleiotropic, affecting both mitotic and meiotic functions. At the nonpermissive temperature of 37 degrees C, lyt1 haploid strains budded at a distal position (instead of an axial one, as in wild-type haploid strains) and underwent autolysis when the buds were almost the size of the mother cells. These mitotic alterations in cell stability and budding topology were dependent on growth and protein synthesis. Autolysis was prevented by inhibiting DNA synthesis (with hydroxyurea) or by blocking the assembly of microtubules (with benomyl), suggesting that loss of cell viability must occur at a fixed mitotic cycle stage after DNA synthesis and mitotic spindle assembly. On the other hand, lyt1-1/lyt1-1 diploids failed to sporulate at both 24 and 37 degrees C. Taking into account these characteristics, the lyt1 mutant could be considered a cdc-like mutant. By genetic transformation of an appropriate lyt1 strain with a genomic library, ligated to the multicopy vector YEp13, we isolated a gene capable of complementing mitotic alterations but not the meiotic defect. This was the sporulation-specific gene SPO12, which is expressed under the control of the locus MAT in meiosis and is also expressed in the mitotic cycle (V. Parkes and L. H. Johnston, Nucleic Acids Res. 20:5617-5623, 1992). A significant level of SPO12 mRNA can be detected when this gene is inserted in a multicopy plasmid.

Base Sequence↗
PubMed1998

The lytE gene of Bacillus subtilis 168 encodes a cell wall hydrolase.

Bacillus subtilis cell wall-bound protein CWBP33 is encoded by lytE, a gene expressed during the exponential growth phase. Sequence analysis of LytE, a 33-kDa protein, reveals two domains. The N-terminal domain contains a threefold-repeated motif common to several peptidoglycan binding proteins, while the C-terminal domain, probably carrying the catalytic activity, has homology with certain exoproteins. Zymographs unambiguously reveal that the absence of CWBP33, due to inactivation of lytE, is accompanied by the loss of a lytic activity. In lytE mutants, the cell autolysis rate is significantly decreased, although autolysis of corresponding, purified cell walls does not seem to be affected.

Amino Acid Sequence↗
PubMed1993

Interference with the oxidative response of neutrophils by Streptococcus pneumoniae.

BACKGROUND: Pneumococcal infections are still a major clinical problem. Polymorphonuclear leucocytes (neutrophils) are considered to have a key role in the host's defence against Streptococcus pneumoniae but the mechanisms by which they kill the pneumococcus remain unclear. As reactive oxygen species are regarded as a major antimicrobial defence of phagocytes an attempt has been made to establish their role in the response of neutrophils to S pneumoniae. METHODS: S pneumoniae isolated from patients with bacteraemic pneumococcal pneumonia were incubated with neutrophils in suspension and superoxide production was measured by reduction of ferricytochrome c. RESULTS: S pneumoniae did not stimulate superoxide production alone or in the presence of normal human serum. Spontaneous superoxide production by neutrophils was actually abrogated by S pneumoniae, as was the powerful respiratory burst stimulated by phorbol myristate acetate. This phenomenon depended on both the dose and the viability of the bacteria. With S pneumoniae in the logarithmic phase of growth inhibitory activity was confined to the organisms themselves but with organisms undergoing autolysis it was also present in filtered supernatants, suggesting that the inhibitory activity can be attributed to a factor released during autolysis. CONCLUSIONS: S pneumoniae can interfere with the respiratory burst of neutrophils. This property may help to explain the pathogenicity of the organism.

Humans↗
PubMed1985

Development of a cell wash buffer that minimizes nucleic acid loss from Clostridium perfringens 10543 A.

Autolytic activity and nucleic loss from Clostridium perfringens 10543 A was demonstrated during successive cell washes in hypotonic TES buffer. Autolysis increased nearly sixfold and nucleic acid loss nearly twofold when 10 mM EDTA was added to 0.3 M Tris-sucrose buffer. Attempts to minimize both autolysis and nucleic acid loss from C. perfringens during routine washing steps were unsuccessful when the effects of sucrose concentration, pH, CaCl2 addition, or wash temperature were examined independently. However, autolytic activity was eliminated and nucleic acid loss reduced to less than 5% when C. perfringens cells were washed at 4 or 25 degrees C in 1.0 M sucrose, 50 mM Tris--HCl, and 25 mM CaCl2 at pH 5.7.

Bacteriological Techniques↗
PubMed1981

Swelling of mitochondria in immersion-fixed liver tissue. Effect of various fixatives and of delayed fixation.

In order to study the occurrence of swollen and disrupted mitochondria in tissue preserved for electron microscopy by ordinary fixation methods, liver tissue from miniature pig fetuses was immersion-fixed in fixatives with various types and concentrations of fixing agents and vehicles. Also commercial and purified products have been tested, different fixation times and temperatures as well as the consequences of a short rinsing in buffer solutions prior to fixation. Furthermore, the significance of delayed fixation (autolysis) was studied. It was found that swollen and disrupted mitochondria occur predominantly in liver cells exposed to low concentrations of glutaraldehyde. It is shown that this phenomenon is a result of a specific effect of glutaraldehyde on the mitochondrial membranes. It is not accompanied by parallel changes of other organelles or nuclei, and it not provoked by other fixing agents, vehicles or by delayed fixation (autolysis).

Animals↗