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Acta Microbiologica Polonica 1996
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International Journal of Molecular... Jul 2021The insect immune response is initiated by the recognition of invading microorganisms. Peptidoglycan recognition proteins (PGRPs) function primarily as pattern...
The insect immune response is initiated by the recognition of invading microorganisms. Peptidoglycan recognition proteins (PGRPs) function primarily as pattern recognition receptors by specifically binding to peptidoglycans expressed on microbial surfaces. We cloned a full-length cDNA for a PGRP from the Asian corn borer (Guenée) and designated it as PGRP1. mRNA was mainly detected in the fat bodies and hemocytes. Its transcript levels increased significantly upon bacterial and fungal challenges. Purified recombinant PGRP1 exhibited binding activity to the gram-positive , gram-negative , entomopathogenic fungi , and yeast The binding further induced their agglutination. Additionally, PGRP1 preferred to bind to Lys-type peptidoglycans rather than DAP-type peptidoglycans. The addition of recombinant PGRP1 to plasma resulted in a significant increase in phenoloxidase activity. The injection of recombinant PGRP1 into larvae led to a significantly increased expression of several antimicrobial peptide genes. Taken together, our results suggest that PGRP1 potentially recognizes the invading microbes and is involved in the immune response in .
Topics: Animals; Beauveria; Fat Body; Hemocytes; Immunity, Innate; Insect Proteins; Lepidoptera; Micrococcus luteus; Monophenol Monooxygenase; Peptidoglycan; Pore Forming Cytotoxic Proteins; Saccharomycetales
PubMed: 34360963
DOI: 10.3390/ijms22158198 -
Journal of Bacteriology Jul 1973Electron microscope examination of negatively stained or thin-sectioned cells of Spirochaeta stenostrepta treated with penicillin or lysozyme showed that the...
Electron microscope examination of negatively stained or thin-sectioned cells of Spirochaeta stenostrepta treated with penicillin or lysozyme showed that the peptidoglycan was present as a thin, electron-dense layer adjacent and external to the cytoplasmic membrane. The peptidoglycan was isolated from cells of S. stenostrepta and Spirochaeta litoralis by a procedure including treatments with sodium lauryl sulfate and Pronase. Hydrolysates of the isolated S. stenostrepta and S. litoralis peptidoglycans contained glucosamine, muramic acid, glutamic acid, l-ornithine, and alanine in molar ratios of 0.90:0.85:1.00:1.00:1.40 and of 0.63:0.63:0.99:1.00:1.41, respectively. Determination of N-terminal residues suggested that nearly 50% of the ornithine in S. stenostrepta and S. litoralis peptidoglycans was involved in peptide cross-linkage. The peptidoglycan layer of S. stenostrepta was sensitive to lysozyme and myxobacter AL-1 protease.
Topics: Alanine; Anaerobiosis; Cell Wall; Chromatography, Paper; Chromatography, Thin Layer; Deoxycholic Acid; Glucosamine; Glutamates; Hydrolysis; Microscopy, Electron; Muramic Acids; Muramidase; Ornithine; Penicillin G; Peptide Hydrolases; Peptidoglycan; Pronase; Sodium Dodecyl Sulfate; Spirochaeta; Staining and Labeling; Stereoisomerism
PubMed: 4123918
DOI: 10.1128/jb.115.1.426-435.1973 -
Journal of Bacteriology Feb 2011Few therapeutic alternatives remain for the treatment of infections due to multiresistant Mycobacterium abscessus. Here we show that the peptidoglycans of the "rough"...
Few therapeutic alternatives remain for the treatment of infections due to multiresistant Mycobacterium abscessus. Here we show that the peptidoglycans of the "rough" and "smooth" morphotypes contain predominantly 3→3 cross-links generated by l,d-transpeptidases, indicating that these enzymes are attractive targets for the development of efficient drugs.
Topics: Bacterial Proteins; Cell Wall; Chromatography, High Pressure Liquid; Mycobacterium; Peptidoglycan; Peptidyl Transferases; Tandem Mass Spectrometry
PubMed: 21097619
DOI: 10.1128/JB.00606-10 -
Analytical and Bioanalytical Chemistry Jan 2017Peptidoglycan (PG) is an essential component of the bacterial cell envelope. This macromolecule consists of glycan chains alternating N-acetylglucosamine and...
Peptidoglycan (PG) is an essential component of the bacterial cell envelope. This macromolecule consists of glycan chains alternating N-acetylglucosamine and N-acetylmuramic acid, cross-linked by short peptides containing nonstandard amino acids. Structural analysis of PG usually involves enzymatic digestion of glycan strands and separation of disaccharide peptides by reversed-phase HPLC followed by collection of individual peaks for MALDI-TOF and/or tandem mass spectrometry. Here, we report a novel strategy using shotgun proteomics techniques for a systematic and unbiased structural analysis of PG using high-resolution mass spectrometry and automated analysis of HCD and ETD fragmentation spectra with the Byonic software. Using the PG of the nosocomial pathogen Clostridium difficile as a proof of concept, we show that this high-throughput approach allows the identification of all PG monomers and dimers previously described, leaving only disambiguation of 3-3 and 4-3 cross-linking as a manual step. Our analysis confirms previous findings that C. difficile peptidoglycans include mainly deacetylated N-acetylglucosamine residues and 3-3 cross-links. The analysis also revealed a number of low abundance muropeptides with peptide sequences not previously reported. Graphical Abstract The bacterial cell envelope includes plasma membrane, peptidoglycan, and surface layer. Peptidoglycan is unique to bacteria and the target of the most important antibiotics; here it is analyzed by mass spectrometry.
Topics: Automation; Bacterial Proteins; Chemistry Techniques, Analytical; Peptidoglycan; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 27520322
DOI: 10.1007/s00216-016-9857-5 -
Proceedings of the National Academy of... Dec 2008The stress-bearing component of the bacterial cell wall--a multi-gigadalton bag-like molecule called the sacculus--is synthesized from peptidoglycan. Whereas the...
The stress-bearing component of the bacterial cell wall--a multi-gigadalton bag-like molecule called the sacculus--is synthesized from peptidoglycan. Whereas the chemical composition and the 3-dimensional structure of the peptidoglycan subunit (in at least one conformation) are known, the architecture of the assembled sacculus is not. Four decades' worth of biochemical and electron microscopy experiments have resulted in two leading 3-D peptidoglycan models: "Layered" and "Scaffold", in which the glycan strands are parallel and perpendicular to the cell surface, respectively. Here we resolved the basic architecture of purified, frozen-hydrated sacculi through electron cryotomography. In the Gram-negative sacculus, a single layer of glycans lie parallel to the cell surface, roughly perpendicular to the long axis of the cell, encircling the cell in a disorganized hoop-like fashion.
Topics: Caulobacter crescentus; Cell Wall; Escherichia coli; Models, Molecular; Peptidoglycan
PubMed: 19033194
DOI: 10.1073/pnas.0808035105 -
Canadian Journal of Microbiology Oct 2012The O-acetylation of peptidoglycan in Gram-negative bacteria occurs specifically at the C-6 hydroxyl group of muramoyl residues. The level of peptidoglycan O-acetylation...
The O-acetylation of peptidoglycan in Gram-negative bacteria occurs specifically at the C-6 hydroxyl group of muramoyl residues. The level of peptidoglycan O-acetylation was found to decrease from 51% to 29% upon differentiation of Proteus mirabilis vegetative cells to swarmers. This decrease was accompanied by a change in the muropeptide composition of the peptidoglycan. In particular, the content of anhydromuropeptides increased, while the amount of Lys-Lys-muropeptides arising from bound lipoprotein decreased. These changes together with a shift in proportion of larger muropeptides suggested a decrease in average chain length of the muropeptides from swarmer cells. Zymography using SDS-PAGE gels containing either O-acetylated or chemically de-O-acetylated peptidoglycan was used to monitor the activity of specific autolysins during the differentiation of vegetative to swarming cells of P. mirabilis. A 43 kDa autolysin with increased specificity for O-acetylated peptidoglycan was detected in vegetative cells, but its activity appeared to decrease as the cells began to differentiate, while the levels of 3 other autolysins with apparent specificity for non-O-acetylated peptidoglycan increased. These changes are discussed in relation to the autolysin profile of the bacteria and the changes in peptidoglycan composition with cell differentiation.
Topics: Acetylation; Cell Differentiation; Electrophoresis, Polyacrylamide Gel; N-Acetylmuramoyl-L-alanine Amidase; Peptidoglycan; Proteus mirabilis
PubMed: 23051614
DOI: 10.1139/w2012-102 -
Insect Biochemistry and Molecular... Sep 2022Peptidoglycan recognition proteins (PGRPs) detect invading bacteria to trigger or modulate immune responses in insects. While these roles are established in Drosophila,...
Peptidoglycan recognition proteins (PGRPs) detect invading bacteria to trigger or modulate immune responses in insects. While these roles are established in Drosophila, functional studies are not yet achieved at the PGRP family level in other insects. To attain this goal, we selected Manduca sexta PGRP12 and five of the nine secreted PGRPs for recombinant expression and biochemical characterization. We cloned PGRP2-5, 12 and 13 cDNAs, produced the proteins in full (PGRP2-5, 13) or in part (PGRP3s, 12e, 13N, 13C) in Sf9 cells, and tested their bindings of two muramyl pentapeptides by surface plasmon resonance, two soluble peptidoglycans by competitive ELISA, and four insoluble peptidoglycans and eight whole bacteria by a pull-down assay. Preferential binding of meso-diaminopimelic acid-peptidoglycans (DAP-PGs) was observed in all the proteins containing a peptidoglycan binding domain and, since PGRP6, 7 and 9 proteins were hardly detected in cell-free hemolymph, the reportoire of PGRPs (including PGRP1 published previously) in M. sexta hemolymph is likely adapted to mainly detect Gram-negative bacteria and certain Gram-positive bacteria with DAP-PGs located on their surface. After incubation with plasma from naïve larvae, PGRP2, 3f, 4, 5, 13f and 13N considerably stimulated prophenoloxidase activation in the absence of a bacterial elicitor. PGRP3s and 12e had much smaller effects. Inclusion of the full-length PGRPs and their regions in the plasma also led to proHP8 activation, supporting their connections to the Toll pathway, since HP8 is a Spӓtzle-1 processing enzyme in M. sexta. Together, these findings raised concerns on the common belief that the Toll-pathway is specific for Gram-positive bacteria in insects.
Topics: Animals; Carrier Proteins; Diaminopimelic Acid; Drosophila; Hemolymph; Insect Proteins; Manduca; Nitrogen Radioisotopes; Peptidoglycan
PubMed: 36007680
DOI: 10.1016/j.ibmb.2022.103827 -
Journal of Molecular Biology Feb 2012Mycobacterium tuberculosis ArfA (Rv0899) is a membrane protein encoded by an operon that is required for supporting bacterial growth in acidic environments. Its...
Mycobacterium tuberculosis ArfA (Rv0899) is a membrane protein encoded by an operon that is required for supporting bacterial growth in acidic environments. Its C-terminal domain (C domain) shares significant sequence homology with the OmpA-like family of peptidoglycan-binding domains, suggesting that its physiological function in acid stress protection may be related to its interaction with the mycobacterial cell wall. Previously, we showed that ArfA forms three independently structured modules, and we reported the structure of its central domain (B domain). Here, we describe the high-resolution structure and dynamics of the C domain, we identify ArfA as a peptidoglycan-binding protein and we elucidate the molecular basis for its specific recognition of diaminopimelate-type peptidoglycan. The C domain of ArfA adopts the characteristic fold of the OmpA-like family. It exhibits pH-dependent conformational dynamics (with significant heterogeneity at neutral pH and a more ordered structure at acidic pH), which could be related to its acid stress response. The C domain associates tightly with polymeric peptidoglycan isolated from M. tuberculosis and also associates with a soluble peptide intermediate of peptidoglycan biosynthesis. This enabled us to characterize the peptidoglycan binding site where five highly conserved ArfA residues, including two key arginines, establish the specificity for diaminopimelate- but not Lys-type peptidoglycan. ArfA is the first peptidoglycan-binding protein to be identified in M. tuberculosis. Its functions in acid stress protection and peptidoglycan binding suggest a link between the acid stress response and the physicochemical properties of the mycobacterial cell wall.
Topics: Bacterial Outer Membrane Proteins; Binding Sites; Crystallography, X-Ray; Diaminopimelic Acid; Hydrogen-Ion Concentration; Models, Molecular; Mycobacterium tuberculosis; Peptidoglycan; Protein Conformation
PubMed: 22206986
DOI: 10.1016/j.jmb.2011.12.030 -
Microbial Drug Resistance (Larchmont,... Jun 2014
Topics: Anti-Bacterial Agents; Bacteria; Cell Wall; Humans; Peptidoglycan
PubMed: 24895897
DOI: 10.1089/mdr.2014.1501