Constituent Of Peptidoglycan Research Articles

Impact of the Deletion of Genes of the Nitrogen Metabolism on Triacylglycerol, Cardiolipin and Actinorhodin Biosynthesis in Streptomyces coelicolor.

Since nitrogen limitation is known to be an important trigger of triacylglycerol (TAG) accumulation in most microorganisms, we first assessed the global lipid content of 21 strains derived from Streptomyces coelicolor M145 deleted for genes involved in nitrogen metabolism. Seven of these strains deleted for genes encoding proteins involved in polyamine (GlnA2/SCO2241, GlnA3/SCO6962, GlnA4/SCO1613), or protein (Pup/SCO1646) degradation, in the regulation of nitrogen metabolism (GlnE/SCO2234 and GlnK/SCO5584), or the global regulator DasR/SCO5231 that controls negatively the degradation of N-acetylglucosamine, a constituent of peptidoglycan, had a higher TAG content than the original strain, whereas five of these strains (except the glnA2 and pup mutants) had a lower cardiolipin (CL) content. The production of the blue polyketide actinorhodin (ACT) was totally abolished in the dasR mutant in both Pi conditions, whereas the deletion of pup, glnA2, glnA3, and glnA4 was correlated with a significant increase in total ACT production, but mainly in Pi limitation. Unexpectedly, ACT production was strongly reduced in the glnA3 mutant in Pi proficiency. Altogether, our data suggest that high TAG and ACT biosynthesis and low CL biosynthesis might all contribute to the lowering of oxidative stress resulting from nitrogen limitation or from other causes.

Bacteria-targeted photothermal therapy for combating drug-resistant bacterial infections.

Photothermal therapy is an ideal non-invasive treatment for bacterial infections. However, if photothermal agents are unable to target bacteria, they can also cause thermal damage to healthy tissue. This study describes the fabrication of a Ti3C2Tx MXene-based photothermal nanobactericide (denoted as MPP) that targets bacteria by modifying MXene nanosheets with polydopamine and the bacterial recognition peptide CAEKA. The polydopamine layer blunts the sharp edges of MXene nanosheets, preventing their damage to normal tissue cells. Furthermore, as a constituent of peptidoglycan, CAEKA can recognize and penetrate the bacterial cell membrane based on similar compatibility. The obtained MPP exhibits superior antibacterial activity and high cytocompatibility compared to the pristine MXene nanosheets. In vivo studies showed that MPP colloidal solution under 808 nm NIR light can effectively treat a subcutaneous abscess caused by multi-drug resistant bacterial infection without adverse effects.

Targeting Imd pathway receptor in Drosophila melanogaster and repurposing of phyto-inhibitors: structural modulation and molecular dynamics

Dysbiosis is a major cause of disease in an individual, generally initiated in the gastrointestinal tract. The gut, also known as the second brain, constitutes a major role in immune signaling. To study the immunity cascade, the Drosophila model was considered targeting the Imd pathway receptor (2F2L) located in the midgut. This receptor further initiates the immune signaling mechanism influenced by bacteria. To inhibit the Imd pathway, the crystal structure of Imd with PDB: 2F2L was considered for the screening of suitable ligand/inhibitor. In light of our previous studies, repurposing of anti-diabetic ligands from the banana plant namely lupeol (LUP), stigmasterol (STI), β-sitosterol (BST) and umbelliferone (UMB) were screened. This study identifies the potential inhibitor along with the tracheal toxin (TCT), a major peptidoglycan constituent of microbes. The molecular docking and molecular dynamics simulation of complexes 2F2L-MLD, 2F2L- CAP, 2F2L-LUP, 2F2L-BST, 2F2L-STI and 2F2L-UMB elucidates the intermolecular interaction into the inhibitory property of ligands. The results of this study infer LUP and UMB as better ligands with high stability and functionality among the screened candidates. This study provides insights into the dysbiosis and its amelioration by plant-derived molecules. The identified drugs (LUP & UMB) will probably act as an inhibitor against microbial dysbiosis and other related pathogenesis (diabetes and diabetic neuropathy). Further, this study will widen avenues in fly biology research and which could be used as a therapeutic model in the rapid, reliable and reproducible screening of phytobiologics in complementary and alternative medicine for various lifestyle associated complications.

What Are the Multi-Omics Mechanisms for Adaptation by Microorganisms to High Alkalinity? A Transcriptomic and Proteomic Study of a Bacillus Strain with Industrial Potential.

Alkaliphilic organisms are among an industrially important class of extremophile microorganisms with the ability to thrive at pH 10-11.5. Microorganisms that exhibit alkaliphilic characteristics are sources of alkali-tolerant enzymes such as proteases, starch degrading enzymes, cellulases, and metabolites such as antibiotics, enzyme inhibitors, siderophores, organic acids, and cholic acid derivatives, which have found various applications in industry for human and environmental health. Yet, multi-omics mechanisms governing adaptation to high alkalinity have been poorly studied. We undertook the present work to understand, as a case study, the alkaliphilic adaptation strategy of the novel microorganism, Bacillus marmarensis DSM 21297, to alkaline conditions using a multi-omics approach that employed transcriptomics and proteomics. As alkalinity increased, bacteria remodeled the peptidoglycan layer by changing peptide moieties along with the peptidoglycan constituents and altered the cell membrane to reduce lipid motility and proton leakiness to adjust intracellular pH. Different transporters also contributed to the maintenance of this pH homeostasis. However, unlike in most well-known alkaliphiles, not only sodium ions but also potassium ions were involved in this process. Interestingly, increased pH has triggered the expression of neither general stress proteins nor gene encoding proteins associated with heat, salt, and nutrient stresses. Only an increase in the expression of oxidative stress related genes was evident. Endospore formation, also a phenomenon closely linked to stress, was unclear. This questioned if high pH was a real stress for B. marmarensis. These new findings, corroborated using the multi-omics approach of the present case study, broaden the knowledge on the mechanisms of alkaliphilic adaptation and might also potentially offer useful departure points for further industrial applications with other microorganisms.

Corynebacterium glutamicum possesses β-N-acetylglucosaminidase

BackgroundIn Gram-positive Corynebacterium glutamicum and other members of the suborder Corynebacterianeae, which includes mycobacteria, cell elongation and peptidoglycan biosynthesis is mainly due to polar growth. C. glutamicum lacks an uptake system for the peptidoglycan constituent N-acetylglucosamine (GlcNAc), but is able to catabolize GlcNAc-6-phosphate. Due to its importance in white biotechnology and in order to ensure more sustainable processes based on non-food renewables and to reduce feedstock costs, C. glutamicum strains have previously been engineered to produce amino acids from GlcNAc. GlcNAc also is a constituent of chitin, but it is unknown if C. glutamicum possesses chitinolytic enzymes.ResultsChitin was shown here not to be growth substrate for C. glutamicum. However, its genome encodes a putative N-acetylglucosaminidase. The nagA2 gene product was active as β-N-acetylglucosaminidase with 0.27 mM 4-nitrophenyl N,N’-diacetyl-β-D-chitobioside as substrate supporting half-maximal activity. NagA2 was secreted into the culture medium when overproduced with TAT and Sec dependent signal peptides, while it remained cytoplasmic when overproduced without signal peptide. Heterologous expression of exochitinase gene chiB from Serratia marcescens resulted in chitinolytic activity and ChiB secretion was enhanced when a signal peptide from C. glutamicum was used. Colloidal chitin did not support growth of a strain secreting exochitinase ChiB and β-N-acetylglucosaminidase NagA2.ConclusionsC. glutamicum possesses β-N-acetylglucosaminidase. In the wild type, β-N-acetylglucosaminidase activity was too low to be detected. However, overproduction of the enzyme fused to TAT or Sec signal peptides led to secretion of active β-N-acetylglucosaminidase. The finding that concomitant secretion of endogenous NagA2 and exochitinase ChiB from S. marcescens did not entail growth with colloidal chitin as sole or combined carbon source, may indicate the requirement for higher or additional enzyme activities such as processive chitinase or endochitinase activities.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0795-3) contains supplementary material, which is available to authorized users.

Insidious pathogen-mimicking properties of nanoparticles in triggering the lectin pathway of the complement system

AbstractThe lectin pathway of the complement system is an integral component of the innate immune system recognizing pathogens through patterns of sugar moieties displayed on their surfaces and neutralizing them through an antibody-independent reaction cascade. Many engineered nanoparticles incite complement through the lectin pathway, but these nanoparticles inherently do not express surface-exposed sugars. However, the projected polymeric surface architecture of nanoparticles may transiently resemble structural motifs of peptidoglycan constituents of pathogens and trigger the lectin pathway. We discuss these issues in relation to nanomedicine design and immune safety.

Crohn's disease: a role of gut microbiota and Nod2 gene polymorphisms in disease pathogenesis.

Crohn's disease is a chronic immune-mediated intestinal inflammation targeted against a yet incompletely defined subset of commensal gut microbiota and occurs on the background of a genetic predisposition under the influence of environmental factors. Genome-wide association studies have identified about 70 genetic risk loci associated with Crohn's disease. The greatest risk for Crohn's disease represent polymorphisms affecting the CARD15 gene encoding nucleotide-binding oligomerization domain 2 (NOD2) which is an intracellular sensor for muramyl dipeptide, a peptidoglycan constituent of bacterial cell wall. The accumulated evidence suggests that gut microbiota represent an essential, perhaps a central factor in the induction and maintaining of Crohn's disease where dysregulation of normal co-evolved homeostatic relationships between intestinal microbiota and host mucosal immune system leads to intestinal inflammation. Taken together, these findings identify Crohn's disease as a syndrome of overlapping phenotypes that involves variable influences of genetic and environmental factors. A deeper understanding of different genetic abnormalities underlying Crohn's disease together with the identification of beneficial and harmful components of gut microbiota and their interactions are essential conditions for the categorization of Crohn's disease patients, which enable us to design more effective, preferably causative, individually tailored therapy.

B. anthracisassociated cardiovascular dysfunction and shock: the potential contribution of both non-toxin and toxin components

The development of cardiovascular dysfunction and shock in patients with invasive Bacillus anthracis infection has a particularly poor prognosis. Growing evidence indicates that several bacterial components likely play important pathogenic roles in this injury. As with other pathogenic Gram-positive bacteria, the B. anthracis cell wall and its peptidoglycan constituent produce a robust inflammatory response with its attendant tissue injury, disseminated intravascular coagulation and shock. However, B. anthracis also produces lethal and edema toxins that both contribute to shock. Growing evidence suggests that lethal toxin, a metalloprotease, can interfere with endothelial barrier function as well as produce myocardial dysfunction. Edema toxin has potent adenyl cyclase activity and may alter endothelial function, as well as produce direct arterial and venous relaxation. Furthermore, both toxins can weaken host defense and promote infection. Finally, B. anthracis produces non-toxin metalloproteases which new studies show can contribute to tissue injury, coagulopathy and shock. In the future, an understanding of the individual pathogenic effects of these different components and their interactions will be important for improving the management of B. anthracis infection and shock.

Peptidoglycan N-acetylglucosamine deacetylation decreases autolysis in Lactococcus lactis

The gene xynD (renamed pgdA) of Lactococcus lactis IL1403 was shown to encode a peptidoglycan N-acetylglucosamine deacetylase. Inactivation of pgdA in L. lactis led to fully acetylated peptidoglycan, whereas cloning of pgdA on a multicopy plasmid vector resulted in an increased degree of peptidoglycan deacetylation, as shown by analysis of peptidoglycan constituent muropeptides. An increased amount of N-unsubstituted glucosamine residues in peptidoglycan resulted in a reduction of the rate of autolysis of L. lactis cells. The activity of the L. lactis major autolysin AcmA was tested on L. lactis cells or peptidoglycan with different degrees of de-N-acetylation. Deacetylated peptidoglycan exhibited decreased susceptibility to AcmA hydrolysis. This reduced susceptibility to AcmA did not result from reduced AcmA binding to peptidoglycan with an increasing degree of de-N-acetylation. In conclusion, enzymic N-acetylglucosamine deacetylation protects peptidoglycan from hydrolysis by the major autolysin AcmA in L. lactis cells, and this leads to decreased cellular autolysis.

Microbial exposure of rural school children, as assessed by levels of N-acetyl-muramic acid in mattress dust, and its association with respiratory health

Background Endotoxin exposure has been shown to be associated with a decreased prevalence of atopic sensitization and symptoms. Yet endotoxin represents only a part of the indoor microbial exposure. Muramic acid, a constituent of peptidoglycan, is present in gram-negative and gram-positive bacteria in the environment and may therefore serve as an additional marker of microbial exposure. Objective To study the factors determining the level of indoor exposure to muramic acid/peptidoglycan, as well as its potential association with respiratory health. Methods In 553 farm and nonfarm school children from Austria, Switzerland, and Germany, mattress dust muramic acid concentrations were determined, and health was assessed by using IgE measurements and questionnaire information. Results The muramic acid concentration was found to be significantly higher in dust from farm children's mattresses than in dust from nonfarm children's mattresses (157 vs 131 ng/mg). Children with higher mattress dust muramic acid concentrations had a significantly lower prevalence of wheezing (odds ratio of highest vs lowest tertile of muramic acid concentration, 0.3; 95% CI, 0.1-0.9), regardless of farming status and endotoxin exposure. The association for asthma was similar, and no association was found with atopic sensitization. Conclusion Next to endotoxin, muramic acid provides us with an independent marker of microbial exposure. Unlike endotoxin, muramic acid was inversely associated with wheezing rather than with atopic sensitization.

Distinct Carbohydrate Recognition Domains of an Invertebrate Defense Molecule Recognize Gram-negative and Gram-positive Bacteria

Coelomic fluid of Eisenia foetida earthworms (Oligochaeta, Annelida) contains a 42-kDa defense molecule named CCF for coelomic cytolytic factor. By binding microbial antigens, namely the O-antigen of lipopolysaccharide (LPS), beta-1,3-glucans, or N,N'-diacetylchitobiose present, respectively, on Gram-negative bacteria or yeast cell walls, CCF triggers the prophenoloxidase activating pathway. We report that CCF recognizes lysozyme-predigested Gram-positive bacteria or the peptidoglycan constituent muramyl dipeptide as well as muramic acid. To identify the pattern recognition domains of CCF, deletion mutants were tested for their ability to reconstitute the prophenoloxidase cascade in E. foetida coelomic fluid depleted of endogenous CCF in the presence of LPS, beta-1,3-glucans, N,N'-diacetylchitobiose, and muramic acid. In addition, affinity chromatography of CCF peptides was performed on immobilized beta-1,3-glucans or N,N'-diacetylchitobiose. We found that the broad specificity of CCF for pathogen-associated molecular patterns results from the presence of two distinct pattern recognition domains. One domain, which shows homology with the polysaccharide and glucanase motifs of beta-1,3-glucanases and invertebrate defense molecules located in the central part of the CCF polypeptide chain, interacts with LPS and beta-1,3-glucans. The C-terminal tryptophan-rich domain mediates interactions of CCF with N,N'-diacetylchitobiose and muramic acid. These data provide evidence for the presence of spatially distinct carbohydrate recognition domains within this invertebrate defense molecule.

Permeability of mycobacterial cell envelopes to sterols: Peptidoglycan as the diffusion barrier

AbstractVancomycin, an inhibitor of peptidoglycan synthesis, depressed the growth of Mycobacterium sp. NRRL B 3805 and MB 3683, but not the β‐sitosterol side chain degradation and androstane derivatives accumulation. As a result, the specific activity (products formed/cell weight unit × h) increased threefold in the peptidoglycan‐deficient cells, indicating faster crossing of the sterol through the cell water barrier.Cell wall preparations: crude cell wall (CCW), purified cell wall (PCW), and peptidoglycan — enriched PCW — residue (PEPCW) were obtained and analysed in order to find a relationship between the vancomycin — induced chemical changes and the permeation rate of the sterol.The amounts of CCW, PCW and PEPCW, produced from 8 g lyophilised control cells were 445, 170 and 28 mg respectively. The respective figures were 176, 61, and 4.8 mg for vancomycin — treated cells. In addition to the lower content of the rigid layer, a distinct shift in the molar ratios of the peptidoglycan constituents: alanine, glutamic, diaminopimelic and muramic acids, and glucosamine was observed under the action of the murein inhibitor. The most significant change was that of muramic acid: diaminopimelic acid molar ratio, the compounds which are markers of glycan strands and tetrapeptides, respectively. In control cells it was approximately 1:1, and increased to 1.34–1.43: 1 in the compared preparation, which indicated a marked decrease in the tetrapeptide moieties crosslinking the main glycan strands. Together with the general lower content of murein, this modification may be responsible for the enhanced sterol permeation through the cell wall.

Incorporation of bacterial peptidoglycan constituents into macrophage lipids during phagocytosis.

It has previously been established that several glycopeptides of peptidoglycan origin are formed as a result of processing of Bacillus subtilis cell walls by the macrophage-like cell line RAW264. Although the formation of these glycopeptides could account for the humoral immune responses characteristic of bacterial peptidoglycans, their formation does not account for the cellular-mediated immune responses observed for water-in-oil emulsions of peptidoglycan or for lipophilic derivatives of glycopeptide fragments thereof. Therefore, the processing of peptidoglycan by macrophages was reexamined to establish whether the lipophilic derivative of any peptidoglycan-derived glycopeptide was formed. The experiments were performed by incubating B. subtilis cell walls radiolabeled in muramic acid, glucosamine, alanine, glutamic acid, and diaminopimelic acid residues in the presence of the macrophage-like cell line RAW264. The crude lipid fraction derived from the macrophages was further fractionated and analyzed, revealing the presence of two lipophilic glycopeptides that contained glucosamine, muramic acid, and alanine of bacterial origin.

Distribution of dipeptides containing d-alanine and characteristics of the formation ability of these dipeptides in japonica type rice plant

Abstract Although most of the amino acids in living organisms consist of L-forms, some D-amino acids are well known as the constituents of peptidoglycan in bacterial cell walls and peptide antibiotics produced by microorganisms (Soda 1977). D-Amino acids are present not only in microorganisms but. also in higher plants. For example, large amounts of D-alanine (D-Ala) were detected in the form of dipeptides in pea seedings (Fukuda et al. 1973) and rice leaf blades (Yamauchi et al. 1979; Manabe et al. 1981). However, their role in these plants is not known.

Elimination of group A streptococcal cell walls from mammalian tissues.

The precise levels in mammalian tissues of rhamnose (a constituent of the group-specific polysaccharide) and muramic acid (a constituent of peptidoglycan) at various times after systemic administration of streptococcal cell walls were noted for the first time. The extreme resistance of the streptococcal cell wall to degradation by mammalian enzymes can now be clearly appreciated. The results help explain the chronic nature of certain inflammatory diseases elicited by streptococcal cell wall debris.

Cell wall-polypeptide complexes in Bacillus subtilis

The cell surface of Bacillus subtilis contains several peptidoglycan-associated polypeptides. Cell walls were labeled with 125I or 35S, and the products were digested with lysozyme. When the digests were chromatographed on Sephacryl S-200, peaks of radioactivity corresponding to molecular weights of 240,000, 125,000, 20,000, 17,000, and 15,000 were observed. The walls solubilized by lysozyme were also subjected to sodium dodecyl sulfate-poly(acrylamide) gel electrophoresis, and radioactive bands corresponding to apparent molecular weights of 24,000, 22,000, and 19,000 were found. Isoelectric focusing of the digests revealed the presence of a component having an isoelectric point of 3.7, and, possibly, of minor components having isoelectric points of 4.7 and 6.1. Proteases, including trypsin, subtilisin, and pronase, removed some of the radioactivity from [ 35S]-labeled walls. Significant proportions of label from [ 35S]walls were solubilized by the peptide-bond-breaking agents cyanogen bromide and N-bromosuccinimide. Small proportions of radioactivity were released from labeled walls by hydroxylamine and trichloroacetic acid. Direct, amino acid analyses of the walls showed the presence of several amino acids not commonly regarded as constituents of peptidoglycan. Cell walls from a protease-deficient mutant, and from a wall preparation enriched in cell poles, contained similar proportions of amino acids. In addition, wall preparations from an autolysin-deficient mutant, and walls from protease hyper-producing strains, contained amino acids that could not be removed by rigorous extraction-procedures. The results suggest that the cell walls of Bacillus subtilis contain tightly, or covalently, bound protein molecules or polypeptides that are refractory to removal by denaturants.

Isolation and Characterization of Structural Components of Bacillus cereus AHU 1356 Cell Walls

From lysozyme digests of the N-acetylated cell walls of Bacillus cereus AHU 1356, a polysaccharide fraction and a teichoic acid fraction were isolated by ion-exchange chromatography and gel chromatography. The former fraction, accounting for 50% of the walls, contained N-acetylglucosamine, N-acetylmannosamine, N-acetylgalactosamine and glucose in a molar ratio of 4:1:1:1 together with a small amount of the peptidoglycan constituents. The latter fraction accounted for 5% of the walls and was composed of N-acetylglucosamine, galactose, glycerol and phosphorus in a molar ratio of 1:1.4:1:1 and a small amount of the peptidoglycan constituents. The molecular weight of the polysaccharide fraction was about 33000 as estimated by gel chromatography. After acid hydrolysis of the polysaccharide fraction, a stoichiometric amount of muramic acid 6-phosphate was detected which was probably involved in the linkage between the polysaccharide and peptidoglycan moieties. The polysaccharide moiety detached from the peptidoglycan components by mild acid hydrolysis was isolated as material with an approximate molecular weight of 28000. Structural studies on the polysaccharide, involving Smith degradation, CrO3 oxidation and proton magnetic resonance spectrometry of the polysaccharide fraction together with the analysis of oligosaccharides obtained from partial acid hydrolysis of the polysaccharide fraction, led to the tentative formulation of the most likely structure of the repeating unit as follows:

Localization of protein A in the bacteria.

Staphylococcus aureus, strain Cowan I, contains 1.7% protein A and its isolated cell walls contain 6.7%.Removal of contaminating cytoplasmic membrane fragments from the cell walls does not decrease the content of protein A. Extraction of cell walls with trichloroacetic acid releases teichoic acid but not protein A. Extractions with high concentrations of LiCl do not remove protein A. Only bacteriolytic enzymes can release the protein.Protein A isolated from bacteria after lysozyme digestion contains peptidoglycan constituents. More effective bacteriolytic enzymes such as lysostaphin release peptidoglycan fragments from this protein A preparation.We conclude from these results that protein A is a cell wall component of S. aureus and is covalently linked to the peptidoglycan structure.