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Suzanne Walker - One of the best experts on this subject based on the ideXlab platform.
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Staphylococcus aureus cell growth and division are regulated by an amidase that trims peptides from uncrosslinked Peptidoglycan
Nature Microbiology, 2020Co-Authors: Truc Do, Daniel Kahne, Mariana G Pinho, Kaitlin Schaefer, Ace George Santiago, Pedro B. Fernandes, Suzanne WalkerAbstract:Bacterial cell wall amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so they can separate. An amidase that cleaves uncrosslinked Peptidoglycan and its regulator are identified here and shown to regulate cell growth, rather than separation. This enzyme regulates the density of Peptidoglycan assembly sites, ensuring coordination between cell expansion and cell division. Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton^ 1 . In Staphylococcus aureus , the Peptidoglycan assembly enzymes relocate during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells^ 2 – 4 . But how Peptidoglycan synthesis is regulated throughout the cell cycle is poorly understood^ 5 , 6 . Here, we used a transposon screen to identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes stem peptides from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so that they can separate^ 7 . However, this amidase controls cell growth. In its absence, Peptidoglycan synthesis becomes spatially dysregulated, which causes cells to grow so large that cell division is defective. We show that the cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given subcellular location. Removal of stem peptides from Peptidoglycan at the cell periphery promotes Peptidoglycan synthase relocation to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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staphylococcus aureus cell growth and division are regulated by an amidase that trims peptides from uncrosslinked Peptidoglycan
Nature microbiology, 2020Co-Authors: Kaitlin Schaefer, Daniel Kahne, Mariana G Pinho, Ace George Santiago, Pedro B. Fernandes, Kathryn A Coe, Suzanne WalkerAbstract:Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton1. In Staphylococcus aureus, the Peptidoglycan assembly enzymes relocate during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells2-4. But how Peptidoglycan synthesis is regulated throughout the cell cycle is poorly understood5,6. Here, we used a transposon screen to identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes stem peptides from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so that they can separate7. However, this amidase controls cell growth. In its absence, Peptidoglycan synthesis becomes spatially dysregulated, which causes cells to grow so large that cell division is defective. We show that the cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given subcellular location. Removal of stem peptides from Peptidoglycan at the cell periphery promotes Peptidoglycan synthase relocation to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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the cell cycle in staphylococcus aureus is regulated by an amidase that controls Peptidoglycan synthesis
bioRxiv, 2019Co-Authors: Kaitlin Schaefer, Daniel Kahne, Mariana G Pinho, Ace George Santiago, Pedro B. Fernandes, Kathryn A Coe, Suzanne WalkerAbstract:Abstract Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton. In Staphylococcus aureus, the enzymes that assemble Peptidoglycan move during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells. But how Peptidoglycan synthesis is regulated throughout the cell cycle is not understood. Here we identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes peptide chains from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Typical amidases act after cell division to hydrolyze Peptidoglycan between daughter cells so they can separate. However, we show that this amidase controls cell growth. In its absence, excess Peptidoglycan synthesis occurs at the cell periphery, causing cells to grow so large that cell division is defective. We show that cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given cellular location. Removal of peptide chains from Peptidoglycan at the cell periphery promotes synthase movement to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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ftsw is a Peptidoglycan polymerase that is activated by its cognate penicillin binding protein
bioRxiv, 2018Co-Authors: Atsushi Taguchi, Daniel Kahne, Thomas G Bernhardt, Michael Welsh, Lindsey S Marmont, Wonsik Lee, Suzanne WalkerAbstract:The Peptidoglycan cell wall is essential for the survival and shape maintenance of bacteria. For decades it was thought that only penicillin-binding proteins (PBPs) effected Peptidoglycan synthesis. Recently, it was shown that RodA, a member of the Rod complex involved in side wall Peptidoglycan synthesis, acts as a Peptidoglycan polymerase. RodA is absent or dispensable in many bacteria that contain a cell wall; however, all of these bacteria have a RodA homologue, FtsW, which is a core member of the divisome complex that is essential for septal cell wall assembly. FtsW was previously proposed flip the Peptidoglycan precursor Lipid II to the peripasm, but we report here that FtsW polymerizes Lipid II. We show that FtsW polymerase activity depends on the presence of the class B PBP (bPBP) that it recruits to the septum. We also demonstrate that the polymerase activity of FtsW is required for its function in vivo. Our findings establish FtsW as a Peptidoglycan polymerase that works with its cognate bPBP to produce septal Peptidoglycan during cell division.
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seds proteins are a widespread family of bacterial cell wall polymerases
Nature, 2016Co-Authors: Alexander J Meeske, Eammon P Riley, William P Robins, Tsuyoshi Uehara, John J Mekalanos, Daniel Kahne, Suzanne Walker, Andrew C Kruse, Thomas G Bernhardt, David Z RudnerAbstract:Elongation of rod-shaped bacteria is mediated by a dynamic Peptidoglycan-synthetizing machinery called the Rod complex. Here we report that, in Bacillus subtilis, this complex is functional in the absence of all known Peptidoglycan polymerases. Cells lacking these enzymes survive by inducing an envelope stress response that increases the expression of RodA, a widely conserved core component of the Rod complex. RodA is a member of the SEDS (shape, elongation, division and sporulation) family of proteins, which have essential but ill-defined roles in cell wall biogenesis during growth, division and sporulation. Our genetic and biochemical analyses indicate that SEDS proteins constitute a family of Peptidoglycan polymerases. Thus, B. subtilis and probably most bacteria use two distinct classes of polymerase to synthesize their exoskeleton. Our findings indicate that SEDS family proteins are core cell wall synthases of the cell elongation and division machinery, and represent attractive targets for antibiotic development. SEDS proteins are core Peptidoglycan polymerases involved in bacterial cell wall elongation and division. It has been generally accepted that the cell wall Peptidoglycans of the bacterial exoskeleton are synthesized by penicillin binding proteins (PBPs) known as class A PBPs. Now, using genetic manipulation, phylogenetic analysis and functional experiments in Bacillus subtilis, David Rudner and colleagues have identified SEDS family proteins as the main Peptidoglycan polymerases more broadly conserved than class A PBPs. Specifically in B. subtilis, they show that the SEDS protein RodA, a widely conserved component of the Rod complex involved in elongation of rod-shaped bacteria, acts with class B PBPs as the core cell wall synthase of the cell elongation and division machinery. The authors conclude that B. subtilis and probably most bacteria use two distinct classes of polymerases to synthesize their exoskeleton. This work also suggests that SEDS family proteins should be attractive targets for antibiotic development.
Daniel Kahne - One of the best experts on this subject based on the ideXlab platform.
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Staphylococcus aureus cell growth and division are regulated by an amidase that trims peptides from uncrosslinked Peptidoglycan
Nature Microbiology, 2020Co-Authors: Truc Do, Daniel Kahne, Mariana G Pinho, Kaitlin Schaefer, Ace George Santiago, Pedro B. Fernandes, Suzanne WalkerAbstract:Bacterial cell wall amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so they can separate. An amidase that cleaves uncrosslinked Peptidoglycan and its regulator are identified here and shown to regulate cell growth, rather than separation. This enzyme regulates the density of Peptidoglycan assembly sites, ensuring coordination between cell expansion and cell division. Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton^ 1 . In Staphylococcus aureus , the Peptidoglycan assembly enzymes relocate during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells^ 2 – 4 . But how Peptidoglycan synthesis is regulated throughout the cell cycle is poorly understood^ 5 , 6 . Here, we used a transposon screen to identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes stem peptides from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so that they can separate^ 7 . However, this amidase controls cell growth. In its absence, Peptidoglycan synthesis becomes spatially dysregulated, which causes cells to grow so large that cell division is defective. We show that the cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given subcellular location. Removal of stem peptides from Peptidoglycan at the cell periphery promotes Peptidoglycan synthase relocation to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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staphylococcus aureus cell growth and division are regulated by an amidase that trims peptides from uncrosslinked Peptidoglycan
Nature microbiology, 2020Co-Authors: Kaitlin Schaefer, Daniel Kahne, Mariana G Pinho, Ace George Santiago, Pedro B. Fernandes, Kathryn A Coe, Suzanne WalkerAbstract:Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton1. In Staphylococcus aureus, the Peptidoglycan assembly enzymes relocate during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells2-4. But how Peptidoglycan synthesis is regulated throughout the cell cycle is poorly understood5,6. Here, we used a transposon screen to identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes stem peptides from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so that they can separate7. However, this amidase controls cell growth. In its absence, Peptidoglycan synthesis becomes spatially dysregulated, which causes cells to grow so large that cell division is defective. We show that the cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given subcellular location. Removal of stem peptides from Peptidoglycan at the cell periphery promotes Peptidoglycan synthase relocation to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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the cell cycle in staphylococcus aureus is regulated by an amidase that controls Peptidoglycan synthesis
bioRxiv, 2019Co-Authors: Kaitlin Schaefer, Daniel Kahne, Mariana G Pinho, Ace George Santiago, Pedro B. Fernandes, Kathryn A Coe, Suzanne WalkerAbstract:Abstract Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton. In Staphylococcus aureus, the enzymes that assemble Peptidoglycan move during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells. But how Peptidoglycan synthesis is regulated throughout the cell cycle is not understood. Here we identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes peptide chains from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Typical amidases act after cell division to hydrolyze Peptidoglycan between daughter cells so they can separate. However, we show that this amidase controls cell growth. In its absence, excess Peptidoglycan synthesis occurs at the cell periphery, causing cells to grow so large that cell division is defective. We show that cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given cellular location. Removal of peptide chains from Peptidoglycan at the cell periphery promotes synthase movement to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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ftsw is a Peptidoglycan polymerase that is functional only in complex with its cognate penicillin binding protein
Nature microbiology, 2019Co-Authors: Atsushi Taguchi, Daniel Kahne, Andrew C Kruse, Thomas G Bernhardt, Michael Welsh, Lindsey S Marmont, Wonsik Lee, Megan SjodtAbstract:The Peptidoglycan cell wall is essential for the survival and morphogenesis of bacteria1. For decades, it was thought that only class A penicillin-binding proteins (PBPs) and related enzymes effected Peptidoglycan synthesis. Recently, it was shown that RodA-a member of the unrelated SEDS protein family-also acts as a Peptidoglycan polymerase2-4. Not all bacteria require RodA for growth; however, its homologue, FtsW, is a core member of the divisome complex that appears to be universally essential for septal cell wall assembly5,6. FtsW was previously proposed to translocate the Peptidoglycan precursor lipid II across the cytoplasmic membrane7,8. Here, we report that purified FtsW polymerizes lipid II into Peptidoglycan, but show that its polymerase activity requires complex formation with its partner class B PBP. We further demonstrate that the polymerase activity of FtsW is required for its function in vivo. Thus, our findings establish FtsW as a Peptidoglycan polymerase that works with its cognate class B PBP to produce septal Peptidoglycan during cell division.
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ftsw is a Peptidoglycan polymerase that is activated by its cognate penicillin binding protein
bioRxiv, 2018Co-Authors: Atsushi Taguchi, Daniel Kahne, Thomas G Bernhardt, Michael Welsh, Lindsey S Marmont, Wonsik Lee, Suzanne WalkerAbstract:The Peptidoglycan cell wall is essential for the survival and shape maintenance of bacteria. For decades it was thought that only penicillin-binding proteins (PBPs) effected Peptidoglycan synthesis. Recently, it was shown that RodA, a member of the Rod complex involved in side wall Peptidoglycan synthesis, acts as a Peptidoglycan polymerase. RodA is absent or dispensable in many bacteria that contain a cell wall; however, all of these bacteria have a RodA homologue, FtsW, which is a core member of the divisome complex that is essential for septal cell wall assembly. FtsW was previously proposed flip the Peptidoglycan precursor Lipid II to the peripasm, but we report here that FtsW polymerizes Lipid II. We show that FtsW polymerase activity depends on the presence of the class B PBP (bPBP) that it recruits to the septum. We also demonstrate that the polymerase activity of FtsW is required for its function in vivo. Our findings establish FtsW as a Peptidoglycan polymerase that works with its cognate bPBP to produce septal Peptidoglycan during cell division.
Stephane Mesnage - One of the best experts on this subject based on the ideXlab platform.
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Two-site recognition of Staphylococcus aureus Peptidoglycan by lysostaphin SH3b
Nature Chemical Biology, 2020Co-Authors: Luz S. Gonzalez-delgado, Hannah Walters-morgan, Bartłomiej Salamaga, Angus J. Robertson, Andrea M. Hounslow, Elżbieta Jagielska, Izabela Sabała, Mike P. Williamson, Andrew L. Lovering, Stephane MesnageAbstract:A structural look at the interaction between the SH3b domain of the Peptidoglycan endopeptidase lysostaphin and the target for its antistaphylococcal activity, Peptidoglycan, reveals a mechanism of bacterial cell wall binding. Lysostaphin is a bacteriolytic enzyme targeting Peptidoglycan, the essential component of the bacterial cell envelope. It displays a very potent and specific activity toward staphylococci, including methicillin-resistant Staphylococcus aureus . Lysostaphin causes rapid cell lysis and disrupts biofilms, and is therefore a therapeutic agent of choice to eradicate staphylococcal infections. The C-terminal SH3b domain of lysostaphin recognizes Peptidoglycans containing a pentaglycine crossbridge and has been proposed to drive the preferential digestion of staphylococcal cell walls. Here we elucidate the molecular mechanism underpinning recognition of staphylococcal Peptidoglycan by the lysostaphin SH3b domain. We show that the pentaglycine crossbridge and the peptide stem are recognized by two independent binding sites located on opposite sides of the SH3b domain, thereby inducing a clustering of SH3b domains. We propose that this unusual binding mechanism allows synergistic and structurally dynamic recognition of S. aureus Peptidoglycan and underpins the potent bacteriolytic activity of this enzyme.
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duf3380 domain from a salmonella phage endolysin shows potent n acetylmuramidase activity
Applied and Environmental Microbiology, 2016Co-Authors: Lorena Rodriguezrubio, Hans Gerstmans, Simon Thorpe, Rob Lavigne, Stephane Mesnage, Yves BriersAbstract:ABSTRACT Bacteriophage-encoded endolysins are highly diverse enzymes that cleave the bacterial Peptidoglycan layer. Current research focuses on their potential applications in medicine, in food conservation, and as biotechnological tools. Despite the wealth of applications relying on the use of endolysin, little is known about the enzymatic properties of these enzymes, especially in the case of endolysins of bacteriophages infecting Gram-negative species. Automated genome annotations therefore remain to be confirmed. Here, we report the biochemical analysis and cleavage site determination of a novel Salmonella bacteriophage endolysin, Gp110, which comprises an uncharacterized domain of unknown function (DUF3380; pfam11860) in its C terminus and shows a higher specific activity (34,240 U/μM) than that of 14 previously characterized endolysins active against Peptidoglycan from Gram-negative bacteria (corresponding to 1.7- to 364-fold higher activity). Gp110 is a modular endolysin with an optimal pH of enzymatic activity of pH 8 and elevated thermal resistance. Reverse-phase high-performance liquid chromatography (RP-HPLC) analysis coupled to mass spectrometry showed that DUF3380 has N-acetylmuramidase (lysozyme) activity cleaving the β-(1,4) glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine residues. Gp110 is active against directly cross-linked Peptidoglycans with various peptide stem compositions, making it an attractive enzyme for developing novel antimicrobial agents. IMPORTANCE We report the functional and biochemical characterization of the Salmonella phage endolysin Gp110. This endolysin has a modular structure with an enzymatically active domain and a cell wall binding domain. The enzymatic activity of this endolysin exceeds that of all other endolysins previously characterized using the same methods. A domain of unknown function (DUF3380) is responsible for this high enzymatic activity. We report that DUF3380 has N-acetylmuramidase activity against directly cross-linked Peptidoglycans with various peptide stem compositions. This experimentally verified activity allows better classification and understanding of the enzymatic activities of endolysins, which mostly are inferred by sequence similarities. Three-dimensional structure predictions for Gp110 suggest a fold that is completely different from that of known structures of enzymes with the same Peptidoglycan cleavage specificity, making this endolysin quite unique. All of these features, combined with increased thermal resistance, make Gp110 an attractive candidate for engineering novel endolysin-based antibacterials.
Waldemar Vollmer - One of the best experts on this subject based on the ideXlab platform.
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Regulation of Peptidoglycan synthesis and remodelling.
Nature Reviews Microbiology, 2020Co-Authors: Alexander J F Egan, Jeff Errington, Waldemar VollmerAbstract:Bacteria surround their cell membrane with a net-like Peptidoglycan layer, called sacculus, to protect the cell from bursting and maintain its cell shape. Sacculus growth during elongation and cell division is mediated by dynamic and transient multiprotein complexes, the elongasome and divisome, respectively. In this Review we present our current understanding of how Peptidoglycan synthases are regulated by multiple and specific interactions with cell morphogenesis proteins that are linked to a dynamic cytoskeletal protein, either the actin-like MreB or the tubulin-like FtsZ. Several Peptidoglycan synthases and hydrolases require activation by outer-membrane-anchored lipoproteins. We also discuss how bacteria achieve robust cell wall growth under different conditions and stresses by maintaining multiple Peptidoglycan enzymes and regulators as well as different Peptidoglycan growth mechanisms, and we present the emerging role of LD-transpeptidases in Peptidoglycan remodelling.
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Peptidoglycan in obligate intracellular bacteria.
Molecular Microbiology, 2017Co-Authors: Christian Otten, Waldemar Vollmer, Matteo Brilli, Patrick H. Viollier, Jeanne SaljeAbstract:Peptidoglycan is the predominant stress-bearing structure in the cell envelope of most bacteria, and also a potent stimulator of the eukaryotic immune system. Obligate intracellular bacteria replicate exclusively within the interior of living cells, an osmotically protected niche. Under these conditions Peptidoglycan is not necessarily needed to maintain the integrity of the bacterial cell. Moreover, the presence of Peptidoglycan puts bacteria at risk of detection and destruction by host Peptidoglycan recognition factors and downstream effectors. This has resulted in a selective pressure and opportunity to reduce the levels of Peptidoglycan. In this review we have analysed the occurrence of genes involved in Peptidoglycan metabolism across the major obligate intracellular bacterial species. From this comparative analysis, we have identified a group of predicted 'Peptidoglycan-intermediate' organisms that includes the Chlamydiae, Orientia tsutsugamushi, Wolbachia and Anaplasma marginale. This grouping is likely to reflect biological differences in their infection cycle compared with Peptidoglycan-negative obligate intracellular bacteria such as Ehrlichia and Anaplasma phagocytophilum, as well as obligate intracellular bacteria with classical Peptidoglycan such as Coxiella, Buchnera and members of the Rickettsia genus. The signature gene set of the Peptidoglycan-intermediate group reveals insights into minimal enzymatic requirements for building a Peptidoglycan-like sacculus and/or division septum.
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Chapter 6 – Peptidoglycan
Molecular Medical Microbiology, 2015Co-Authors: Waldemar VollmerAbstract:Peptidoglycan is an essential component of the bacterial cell envelope and protects the cell from bursting due to turgor and maintains cell shape. Composed of glycan chains connected by short peptides, Peptidoglycan forms a net-like macromolecule around the cytoplasmic membrane. There is significant structural variation in the Peptidoglycans of different bacteria. Pathogens modify the Peptidoglycan to become resistant to lysozyme. Peptidoglycan carries covalently attached cell surface components like teichoic acid, capsular polysaccharide and cell wall proteins. Peptidoglycan precursors are synthesized in the cytoplasm and linked to a polyprenyl phosphate lipid for transport across the cytoplasmic membrane. Presumably, Peptidoglycan synthases and hydrolases form dynamic multi-enzyme complexes which polymerize new Peptidoglycan and insert it into the existing cell wall, concomitant with the release of old material. The Peptidoglycan synthesis complexes are controlled by components of the bacterial cytoskeleton. Gram-negative bacteria also regulate Peptidoglycan synthesis by outer-membrane proteins.
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Toward the characterization of Peptidoglycan structure and protein-Peptidoglycan interactions by solid-state NMR spectroscopy.
Journal of the American Chemical Society, 2008Co-Authors: Thomas Kern, Waldemar Vollmer, Bernard Joris, Sabine Hediger, Patrick Müller, Cécile Giustini, Catherine M. Bougault, Jean-pierre SimorreAbstract:Solid-state NMR spectroscopy is applied to intact Peptidoglycan sacculi of the Gram-negative bacterium Escherichia coli. High-quality solid-state NMR spectra allow atom-resolved investigation of the Peptidoglycan structure and dynamics as well as the study of protein-Peptidoglycan interactions.
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Bacterial Peptidoglycan (murein) hydrolases.
Fems Microbiology Reviews, 2008Co-Authors: Waldemar Vollmer, Bernard Joris, Paulette Charlier, Simon J. FosterAbstract:Most bacteria have multiple Peptidoglycan hydrolases capable of cleaving covalent bonds in Peptidoglycan sacculi or its fragments. An overview of the different classes of Peptidoglycan hydrolases and their cleavage sites is provided. The physiological functions of these enzymes include the regulation of cell wall growth, the turnover of Peptidoglycan during growth, the separation of daughter cells during cell division and autolysis. Specialized hydrolases enlarge the pores in the Peptidoglycan for the assembly of large trans-envelope complexes (pili, flagella, secretion systems), or they specifically cleave Peptidoglycan during sporulation or spore germination. Moreover, Peptidoglycan hydrolases are involved in lysis phenomena such as fratricide or developmental lysis occurring in bacterial populations. We will also review the current view on the regulation of autolysins and on the role of cytoplasm hydrolases in Peptidoglycan recycling and induction of β-lactamase.
Mariana G Pinho - One of the best experts on this subject based on the ideXlab platform.
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Staphylococcus aureus cell growth and division are regulated by an amidase that trims peptides from uncrosslinked Peptidoglycan
Nature Microbiology, 2020Co-Authors: Truc Do, Daniel Kahne, Mariana G Pinho, Kaitlin Schaefer, Ace George Santiago, Pedro B. Fernandes, Suzanne WalkerAbstract:Bacterial cell wall amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so they can separate. An amidase that cleaves uncrosslinked Peptidoglycan and its regulator are identified here and shown to regulate cell growth, rather than separation. This enzyme regulates the density of Peptidoglycan assembly sites, ensuring coordination between cell expansion and cell division. Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton^ 1 . In Staphylococcus aureus , the Peptidoglycan assembly enzymes relocate during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells^ 2 – 4 . But how Peptidoglycan synthesis is regulated throughout the cell cycle is poorly understood^ 5 , 6 . Here, we used a transposon screen to identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes stem peptides from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so that they can separate^ 7 . However, this amidase controls cell growth. In its absence, Peptidoglycan synthesis becomes spatially dysregulated, which causes cells to grow so large that cell division is defective. We show that the cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given subcellular location. Removal of stem peptides from Peptidoglycan at the cell periphery promotes Peptidoglycan synthase relocation to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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staphylococcus aureus cell growth and division are regulated by an amidase that trims peptides from uncrosslinked Peptidoglycan
Nature microbiology, 2020Co-Authors: Kaitlin Schaefer, Daniel Kahne, Mariana G Pinho, Ace George Santiago, Pedro B. Fernandes, Kathryn A Coe, Suzanne WalkerAbstract:Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton1. In Staphylococcus aureus, the Peptidoglycan assembly enzymes relocate during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells2-4. But how Peptidoglycan synthesis is regulated throughout the cell cycle is poorly understood5,6. Here, we used a transposon screen to identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes stem peptides from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Amidases typically hydrolyse crosslinked Peptidoglycan between daughter cells so that they can separate7. However, this amidase controls cell growth. In its absence, Peptidoglycan synthesis becomes spatially dysregulated, which causes cells to grow so large that cell division is defective. We show that the cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given subcellular location. Removal of stem peptides from Peptidoglycan at the cell periphery promotes Peptidoglycan synthase relocation to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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the cell cycle in staphylococcus aureus is regulated by an amidase that controls Peptidoglycan synthesis
bioRxiv, 2019Co-Authors: Kaitlin Schaefer, Daniel Kahne, Mariana G Pinho, Ace George Santiago, Pedro B. Fernandes, Kathryn A Coe, Suzanne WalkerAbstract:Abstract Bacteria are protected by a polymer of Peptidoglycan that serves as an exoskeleton. In Staphylococcus aureus, the enzymes that assemble Peptidoglycan move during the cell cycle from the periphery, where they are active during growth, to the division site where they build the partition between daughter cells. But how Peptidoglycan synthesis is regulated throughout the cell cycle is not understood. Here we identify a membrane protein complex that spatially regulates S. aureus Peptidoglycan synthesis. This complex consists of an amidase that removes peptide chains from uncrosslinked Peptidoglycan and a partner protein that controls its activity. Typical amidases act after cell division to hydrolyze Peptidoglycan between daughter cells so they can separate. However, we show that this amidase controls cell growth. In its absence, excess Peptidoglycan synthesis occurs at the cell periphery, causing cells to grow so large that cell division is defective. We show that cell growth and division defects due to loss of this amidase can be mitigated by attenuating the polymerase activity of the major S. aureus Peptidoglycan synthase. Our findings lead to a model wherein the amidase complex regulates the density of Peptidoglycan assembly sites to control Peptidoglycan synthase activity at a given cellular location. Removal of peptide chains from Peptidoglycan at the cell periphery promotes synthase movement to midcell during cell division. This mechanism ensures that cell expansion is properly coordinated with cell division.
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inactivated pbp4 in highly glycopeptide resistant laboratory mutants of staphylococcus aureus
Journal of Biological Chemistry, 1999Co-Authors: Krzysztof Sieradzki, Mariana G Pinho, Alexander TomaszAbstract:Abstract Both vancomycin- and teicoplanin-resistant laboratory mutants of Staphylococcus aureus produce Peptidoglycans of altered composition in which the proportion of highly cross-linked muropeptide species is drastically reduced with a parallel increase in the representation of muropeptide monomers and dimers (Sieradzki, K., and Tomasz, A. (1997) J. Bacteriol. 179, 2557–2566; and Sieradzki, K., and Tomasz, A. (1998) Microb. Drug Resist. 4, 159–168). We now report that the distorted Peptidoglycan composition is related to defects in penicillin-binding protein 4 (PBP4); no PBP4 was detectable by the fluorographic assay in membrane preparations from the mutants, and comparison of the sequence of pbp4 amplified from the mutants indicated disruption of the gene by two types of abnormalities, a 17-amino acid long duplication starting at position 305 of thepbp4 gene was detected in the vancomycin-resistant mutant, and a stop codon was found to be introduced into the pbp4 KTG motif at position 261 in the mutant selected for teicoplanin resistance. Additional common patterns of disturbances in the Peptidoglycan metabolism of the mutants are indicated by the increased sensitivity of mutant cell walls to the M1 muramidase and decreased sensitivity to lysostaphin, which is a reversal of the susceptibility pattern of the parental cell walls. Furthermore, the results of high performance liquid chromatography analysis of lysostaphin digests of Peptidoglycan suggest an increase in the average chain length of the glycan strands in the Peptidoglycan of the glycopeptide-resistant mutants. The increased molar proportion of muropeptide monomers in the cell wall of the glycopeptide-resistant mutants should provide binding sites for the “capture” of vancomycin and teicoplanin molecules, which may be part of the mechanism of glycopeptide resistance inS. aureus.
