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MBio May 2021Gram-negative bacteria have a unique cell envelope with a lipopolysaccharide-containing outer membrane that is tightly connected to a thin layer of peptidoglycan. The...
Gram-negative bacteria have a unique cell envelope with a lipopolysaccharide-containing outer membrane that is tightly connected to a thin layer of peptidoglycan. The tight connection between the outer membrane and peptidoglycan is needed to maintain the outer membrane as an impermeable barrier for many toxic molecules and antibiotics. such as covalently attach the abundant outer membrane-anchored lipoprotein Lpp (Braun's lipoprotein) to tripeptides in peptidoglycan, mediated by the transpeptidases LdtA, LdtB, and LdtC. LdtD and LdtE are members of the same family of ld-transpeptidases but they catalyze a different reaction, the formation of 3-3 cross-links in the peptidoglycan. The function of the sixth homologue in , LdtF, remains unclear, although it has been shown to become essential in cells with inhibited lipopolysaccharide export to the outer membrane. We now show that LdtF hydrolyzes the Lpp-peptidoglycan linkage, detaching Lpp from peptidoglycan, and have renamed LdtF to peptidoglycan -iaminopimelic acid rotein midase (DpaA). We show that the detachment of Lpp from peptidoglycan is beneficial for the cell under certain stress conditions and that the deletion of allows frequent transposon inactivation in the () gene, whose product downregulates lipopolysaccharide biosynthesis. DpaA-like proteins have characteristic sequence motifs and are present in many Gram-negative bacteria, of which some have no Lpp, raising the possibility that DpaA has other substrates in these species. Overall, our data show that the Lpp-peptidoglycan linkage in is more dynamic than previously appreciated. Gram-negative bacteria have a complex cell envelope with two membranes and a periplasm containing the peptidoglycan layer. The outer membrane is firmly connected to the peptidoglycan by highly abundant proteins. The outer membrane-anchored Braun's lipoprotein (Lpp) is the most abundant protein in , and about one-third of the Lpp molecules become covalently attached to tripeptides in peptidoglycan. The attachment of Lpp to peptidoglycan stabilizes the cell envelope and is crucial for the outer membrane to function as a permeability barrier for a range of toxic molecules and antibiotics. So far, the attachment of Lpp to peptidoglycan has been considered to be irreversible. We have now identified an amidase, DpaA, which is capable of detaching Lpp from peptidoglycan, and we show that the detachment of Lpp is important under certain stress conditions. DpaA-like proteins are present in many Gram-negative bacteria and may have different substrates in these species.
Topics: Amidohydrolases; Diaminopimelic Acid; Escherichia coli; Escherichia coli Proteins; Lipoproteins; Peptidoglycan
PubMed: 33947763
DOI: 10.1128/mBio.00836-21 -
FEMS Microbiology Reviews Jan 2021Microbiota, and the plethora of signalling molecules that they generate, are a major driving force that underlies a striking range of inter-individual physioanatomic and... (Review)
Review
Microbiota, and the plethora of signalling molecules that they generate, are a major driving force that underlies a striking range of inter-individual physioanatomic and behavioural consequences for the host organism. Among the bacterial effectors, one finds peptidoglycan, the major constituent of the bacterial cell surface. In the steady-state, fragments of peptidoglycan are constitutively liberated from bacterial members of the gut microbiota, cross the gut epithelial barrier and enter the host system. The fate of these peptidoglycan fragments, and the outcome for the host, depends on the molecular nature of the peptidoglycan, as well the cellular profile of the recipient tissue, mechanism of cell entry, the expression of specific processing and recognition mechanisms by the cell, and the local immune context. At the target level, physiological processes modulated by peptidoglycan are extremely diverse, ranging from immune activation to small molecule metabolism, autophagy and apoptosis. In this review, we bring together a fragmented body of literature on the kinetics and dynamics of peptidoglycan interactions with the mammalian host, explaining how peptidoglycan functions as a signalling molecule in the host under physiological conditions, how it disseminates within the host, and the cellular responses to peptidoglycan.
Topics: Animals; Bacteria; Host-Pathogen Interactions; Humans; Mammals; Peptidoglycan; Signal Transduction
PubMed: 32897324
DOI: 10.1093/femsre/fuaa044 -
Proceedings of the National Academy of... Jun 2023Peptidoglycan is a critical component of the bacteria cell envelope. Remodeling of the peptidoglycan is required for numerous essential cellular processes and has been...
Peptidoglycan is a critical component of the bacteria cell envelope. Remodeling of the peptidoglycan is required for numerous essential cellular processes and has been linked to bacterial pathogenesis. Peptidoglycan deacetylases that remove the acetyl group of the -acetylglucosamine (NAG) subunit protect bacterial pathogens from immune recognition and digestive enzymes secreted at the site of infection. However, the full extent of this modification on bacterial physiology and pathogenesis is not known. Here, we identify a polysaccharide deacetylase of the intracellular bacterial pathogen and define a two-tiered role for this enzyme in pathogenesis. First, NAG deacetylation is important for the proper localization and function of the Type IVb secretion system, linking peptidoglycan editing to the modulation of host cellular processes through the action of secreted virulence factors. As a consequence, the vacuole mis-traffics along the endocytic pathway to the lysosome, preventing the formation of a replication permissive compartment. Second, within the lysosome, the inability to deacetylate the peptidoglycan renders the bacteria more sensitive to lysozyme-mediated degradation, resulting in increased bacterial death. Thus, the ability to deacetylate NAG is important for bacteria to persist within host cells and in turn, virulence. Collectively, these results expand the function of peptidoglycan deacetylases in bacteria, linking peptidoglycan editing, Type IV secretion, and the intracellular fate of a bacterial pathogen.
Topics: Humans; Legionella pneumophila; Peptidoglycan; Vacuoles; Legionella; Lysosomes; Bacterial Proteins; Legionnaires' Disease
PubMed: 37252954
DOI: 10.1073/pnas.2119658120 -
Molecular Microbiology Jul 2021Until recently, class A penicillin-binding proteins (aPBPs) were the only enzymes known to catalyze glycan chain polymerization from lipid II in bacteria. Hence, the... (Review)
Review
Until recently, class A penicillin-binding proteins (aPBPs) were the only enzymes known to catalyze glycan chain polymerization from lipid II in bacteria. Hence, the discovery of two novel lipid II polymerases, FtsW and RodA, raises new questions and has consequently received a lot of attention from the research community. FtsW and RodA are essential and highly conserved members of the divisome and elongasome, respectively, and work in conjunction with their cognate class B PBPs (bPBPs) to synthesize the division septum and insert new peptidoglycan into the lateral cell wall. The identification of FtsW and RodA as peptidoglycan glycosyltransferases has raised questions regarding the role of aPBPs in peptidoglycan synthesis and fundamentally changed our understanding of the process. Despite their dethronement, aPBPs are essential in most bacteria. So, what is their function? In this review, we discuss recent progress in answering this question and present our own views on the topic.
Topics: Bacillus subtilis; Bacterial Proteins; Cell Wall; Escherichia coli; Escherichia coli Proteins; Membrane Proteins; Penicillin-Binding Proteins; Peptidoglycan; Peptidoglycan Glycosyltransferase; Staphylococcus aureus; Streptococcus pneumoniae; Uridine Diphosphate N-Acetylmuramic Acid
PubMed: 33709487
DOI: 10.1111/mmi.14714 -
Journal of Bacteriology Mar 2023The dynamic composition of the peptidoglycan cell wall has been the subject of intense research for decades, yet how bacteria coordinate the synthesis of new... (Review)
Review
The dynamic composition of the peptidoglycan cell wall has been the subject of intense research for decades, yet how bacteria coordinate the synthesis of new peptidoglycan with the turnover and remodeling of existing peptidoglycan remains elusive. Diversity and redundancy within peptidoglycan synthases and peptidoglycan autolysins, enzymes that degrade peptidoglycan, have often made it challenging to assign physiological roles to individual enzymes and determine how those activities are regulated. For these reasons, peptidoglycan glycosidases, which cleave within the glycan strands of peptidoglycan, have proven veritable masters of misdirection over the years. Unlike many of the broadly conserved peptidoglycan synthetic complexes, diverse bacteria can employ unrelated glycosidases to achieve the same physiological outcome. Additionally, although the mechanisms of action for many individual enzymes have been characterized, apparent conserved homologs in other organisms can exhibit an entirely different biochemistry. This flexibility has been recently demonstrated in the context of three functions critical to vegetative growth: (i) release of newly synthesized peptidoglycan strands from their membrane anchors, (ii) processing of peptidoglycan turned over during cell wall expansion, and (iii) removal of peptidoglycan fragments that interfere with daughter cell separation during cell division. Finally, the regulation of glycosidase activity during these cell processes may be a cumulation of many factors, including protein-protein interactions, intrinsic substrate preferences, substrate availability, and subcellular localization. Understanding the true scope of peptidoglycan glycosidase activity will require the exploration of enzymes from diverse organisms with equally diverse growth and division strategies.
Topics: Glycoside Hydrolases; Peptidoglycan; Bacteria; Cell Wall; Cell Division; Bacterial Proteins
PubMed: 36757204
DOI: 10.1128/jb.00428-22 -
Annual Review of Biochemistry Jun 2022Biosynthesis of many important polysaccharides (including peptidoglycan, lipopolysaccharide, and -linked glycans) necessitates the transport of lipid-linked... (Review)
Review
Biosynthesis of many important polysaccharides (including peptidoglycan, lipopolysaccharide, and -linked glycans) necessitates the transport of lipid-linked oligosaccharides (LLO) across membranes from their cytosolic site of synthesis to their sites of utilization. Much of our current understanding of LLO transport comes from genetic, biochemical, and structural studies of the multidrug/oligosaccharidyl-lipid/polysaccharide (MOP) superfamily protein MurJ, which flips the peptidoglycan precursor lipid II. MurJ plays a pivotal role in bacterial cell wall synthesis and is an emerging antibiotic target. Here, we review the mechanism of LLO flipping by MurJ, including the structural basis for lipid II flipping and ion coupling. We then discuss inhibition of MurJ by antibacterials, including humimycins and the phage M lysis protein, as well as how studies on MurJ could provide insight into other flippases, both within and beyond the MOP superfamily.
Topics: Bacteria; Lipids; Peptidoglycan; Phospholipid Transfer Proteins
PubMed: 35320686
DOI: 10.1146/annurev-biochem-040320-105145 -
ELife Feb 2021Many antibiotics target the assembly of cell wall peptidoglycan, an essential, heteropolymeric mesh that encases most bacteria. In rod-shaped bacteria, cell wall...
Many antibiotics target the assembly of cell wall peptidoglycan, an essential, heteropolymeric mesh that encases most bacteria. In rod-shaped bacteria, cell wall elongation is spatially precise yet relies on limited pools of lipid-linked precursors that generate and are attracted to membrane disorder. By tracking enzymes, substrates, and products of peptidoglycan biosynthesis in , we show that precursors are made in plasma membrane domains that are laterally and biochemically distinct from sites of cell wall assembly. Membrane partitioning likely contributes to robust, orderly peptidoglycan synthesis, suggesting that these domains help template peptidoglycan synthesis. The cell wall-organizing protein DivIVA and the cell wall itself promote domain homeostasis. These data support a model in which the peptidoglycan polymer feeds back on its membrane template to maintain an environment conducive to directional synthesis. Our findings are applicable to rod-shaped bacteria that are phylogenetically distant from , indicating that horizontal compartmentalization of precursors may be a general feature of bacillary cell wall biogenesis.
Topics: Cell Cycle; Cell Membrane; Cell Wall; Mycobacterium smegmatis; Peptidoglycan
PubMed: 33544079
DOI: 10.7554/eLife.60263 -
Proceedings of the National Academy of... Aug 2023The cell envelope of Gram-negative bacteria consists of three distinct layers: the cytoplasmic membrane, a cell wall made of peptidoglycan (PG), and an asymmetric outer...
The cell envelope of Gram-negative bacteria consists of three distinct layers: the cytoplasmic membrane, a cell wall made of peptidoglycan (PG), and an asymmetric outer membrane (OM) composed of phospholipid in the inner leaflet and lipopolysaccharide (LPS) glycolipid in the outer leaflet. The PG layer has long been thought to be the major structural component of the envelope protecting cells from osmotic lysis and providing them with their characteristic shape. In recent years, the OM has also been shown to be a load-bearing layer of the cell surface that fortifies cells against internal turgor pressure. However, whether the OM also plays a role in morphogenesis has remained unclear. Here, we report that changes in LPS synthesis or modification predicted to strengthen the OM can suppress the growth and shape defects of mutants with reduced activity in a conserved PG synthesis machine called the Rod complex (elongasome) that is responsible for cell elongation and shape determination. Evidence is presented that OM fortification in the shape mutants restores the ability of MreB cytoskeletal filaments to properly orient the synthesis of new cell wall material by the Rod complex. Our results are therefore consistent with a role for the OM in the propagation of rod shape during growth in addition to its well-known function as a diffusion barrier promoting the intrinsic antibiotic resistance of Gram-negative bacteria.
Topics: Lipopolysaccharides; Cell Wall; Cell Membrane; Cytoskeleton; Cell Cycle; Escherichia coli; Peptidoglycan
PubMed: 37607228
DOI: 10.1073/pnas.2301987120 -
ELife May 2020Profiling the phenotype of 200,000 mutants revealed a new cofactor that may help a group of rod-shaped bacteria elongate and grow.
Profiling the phenotype of 200,000 mutants revealed a new cofactor that may help a group of rod-shaped bacteria elongate and grow.
Topics: Bacteria; Cell Wall; Peptidoglycan; Phenotype
PubMed: 32379045
DOI: 10.7554/eLife.57043 -
International Journal of Molecular... Nov 2021Bacteria must maintain the ability to modify and repair the peptidoglycan layer without jeopardising its essential functions in cell shape, cellular integrity and... (Review)
Review
Bacteria must maintain the ability to modify and repair the peptidoglycan layer without jeopardising its essential functions in cell shape, cellular integrity and intermolecular interactions. A range of new experimental techniques is bringing an advanced understanding of how bacteria regulate and achieve peptidoglycan synthesis, particularly in respect of the central role played by complexes of Sporulation, Elongation or Division (SEDs) and class B penicillin-binding proteins required for cell division, growth and shape. In this review we highlight relationships implicated by a bioinformatic approach between the outer membrane, cytoskeletal components, periplasmic control proteins, and cell elongation/division proteins to provide further perspective on the interactions of these cell division, growth and shape complexes. We detail the network of protein interactions that assist in the formation of peptidoglycan and highlight the increasingly dynamic and connected set of protein machinery and macrostructures that assist in creating the cell envelope layers in Gram-negative bacteria.
Topics: Bacterial Proteins; Cell Membrane; Cell Wall; Gram-Negative Bacteria; Penicillin-Binding Proteins; Peptidoglycan; Periplasmic Proteins
PubMed: 34884635
DOI: 10.3390/ijms222312831