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Concept: Peptidoglycan


Lysozymes are key effectors of the animal innate immunity system that kill bacteria by hydrolyzing peptidoglycan, their major cell wall constituent. Recently, specific inhibitors of the three major lysozyme families occuring in the animal kingdom (c-, g- and i-type) have been discovered in Gram-negative bacteria, and it has been proposed that these may help bacteria to evade lysozyme mediated lysis during interaction with an animal host. Escherichia coli produces two inhibitors that are specific for c-type lysozyme (Ivy, Inhibitor of vertebrate lysozyme; MliC, membrane bound lysozyme inhibitor of c-type lysozyme), and one specific for g-type lysozyme (PliG, periplasmic lysozyme inhibitor of g-type lysozyme). Here, we investigated the role of these lysozyme inhibitors in virulence of Avian Pathogenic E. coli (APEC) using a serum resistance test and a subcutaneous chicken infection model. Knock-out of mliC caused a strong reduction in serum resistance and in in vivo virulence that could be fully restored by genetic complementation, whereas ivy and pliG could be knocked out without effect on serum resistance and virulence. This is the first in vivo evidence for the involvement of lysozyme inhibitors in bacterial virulence. Remarkably, the virulence of a ivy mliC double knock-out strain was restored to almost wild-type level, and this strain also had a substantial residual periplasmic lysozyme inhibitory activity that was higher than that of the single knock-out strains. This suggests the existence of an additional periplasmic lysozyme inhibitor in this strain, and indicates a regulatory interaction in the expression of the different inhibitors.

Concepts: Immune system, Archaea, Bacteria, Microbiology, Virus, Escherichia coli, Escherichia coli O157:H7, Peptidoglycan


ABSTRACT Viscoelastic deformation of the contact volume between adhering bacteria and substratum surfaces plays a role in their adhesion and detachment. Currently, there are no deformation models that account for the heterogeneous structure and composition of bacteria, consisting of a relatively soft outer layer and a more rigid, hard core enveloped by a cross-linked peptidoglycan layer. The aim of this paper is to present a new, simple model to derive the reduced Young’s modulus of the contact volume between adhering bacteria and substratum surfaces based on the relationship between deformation and applied external loading force, measured using atomic force microscopy. The model assumes that contact is established through a cylinder with constant volume and does not require assumptions on the properties and dimensions of the contact cylinder. The reduced Young’s moduli obtained (8 to 47 kPa) and dimensions of the contact cylinders could be interpreted on the basis of the cell surface features and cell wall characteristics, i.e., surfaces that are more rigid (because of either less fibrillation, less extracellular polymeric substance production, or a higher degree of cross-linking of the peptidoglycan layer) had shorter contact cylinders and higher reduced Young’s moduli. Application of an existing Hertz model to our experimental data yielded reduced Young’s moduli that were up to 100 times higher for all strains investigated, likely because the Hertz model pertains to a major extent to the more rigid peptidoglycan layer and not only to the soft outer bacterial cell surface, involved in the bond between a bacterium and a substratum surface. IMPORTANCE The viscoelastic properties of the bond between an adhering bacterium and a substratum surface play a role in determining bacterial detachment. For instance, removal of an oral biofilm proceeds according to a viscoelastic failure model, and biofilm left behind after toothbrushing has been found to possess expanded bond lengths between adhering bacteria due to viscoelastic deformation. Current elastic deformation models are unable to distinguish between the soft outer bacterial cell surface and the hard core of a bacterium, enveloped by a peptidoglycan layer. Therefore, here we present a simple model to calculate the Young’s modulus and deformation of the contact volume between an adhering bacterium and a substratum surface that accounts for the heterogeneous structure of a bacterium.

Concepts: Archaea, Bacteria, Cell membrane, Cell wall, Bacterial cell structure, Peptidoglycan, Young's modulus, Deformation


Peptidoglycan is the major structural component of the bacterial cell wall. It provides resistance against turgor and its cleavage by hydrolases such as lysozymes results in bacteriolysis. Most, if not all, animals produce lysozymes as key effectors of their innate immune system. Recently, highly specific bacterial proteinaceous lysozyme inhibitors against the three major animal lysozyme families have been discovered in bacteria, and these may represent a bacterial answer to animal lysozymes. Here, we will review their properties and phylogenetic distribution, present their structure and molecular interaction mechanism with lysozyme, and discuss their possible biological functions and potential applications.

Concepts: Immune system, Archaea, Bacteria, Molecular biology, Enzyme, Organism, Cell wall, Peptidoglycan


Lactobacillus helveticus is traditionally used in dairy industry as a starter or an adjunct culture for manufacture of cheese and some types of fermented milk. Its autolysis releases intracellular enzymes which is a prerequisite for optimum cheese maturation, and is known to be strain dependent. Autolysis is caused by an enzymatic hydrolysis of the cell wall peptidoglycan (PG) by endogenous peptidoglycan hydrolases (PGHs) or autolysins. Origins of differences in autolytic properties of different strains are not fully elucidated. Regulation of autolysis possibly depends on the structure of the cell wall components other than PG, particularly polysaccharides. In the present work, we screened six L. helveticus strains with different autolytic properties: DPC4571, BROI and LH1. We established, for the first time, that cell walls (CWs) of these strains contained polysaccharides, different from their CW teichoic acids. Cell wall polysaccharides of three strains were purified, and their chemical structures were established by 2D NMR spectroscopy and methylation analysis. The structures of their repeating units are presented.

Concepts: Cell, Archaea, Bacteria, Enzyme, Milk, Cell wall, Peptidoglycan, Pseudopeptidoglycan


The distal colon functions as a bioreactor and harbors an enormous amount of bacteria in a mutualistic relationship with the host. The microbiota have to be kept at a safe distance to prevent inflammation, something that is achieved by a dense inner mucus layer that lines the epithelial cells. The large polymeric nets made up by the heavily O-glycosylated MUC2 mucin forms this physical barrier. Proteomic analyses of mucus have identified the lectin-like protein ZG16 (zymogen granulae protein 16) as an abundant mucus component. To elucidate the function of ZG16, we generated recombinant ZG16 and studied Zg16(-/-) mice. ZG16 bound to and aggregated Gram-positive bacteria via binding to the bacterial cell wall peptidoglycan. Zg16(-/-) mice have a distal colon mucus layer with normal thickness, but with bacteria closer to the epithelium. Using distal colon explants mounted in a horizontal perfusion chamber we demonstrated that treatment of bacteria with recombinant ZG16 hindered bacterial penetration into the mucus. The inner colon mucus of Zg16(-/-) animals had a higher load of Gram-positive bacteria and showed bacteria with higher motility in the mucus close to the host epithelium compared with cohoused littermate Zg16(+/+) The more penetrable Zg16(-/-) mucus allowed Gram-positive bacteria to translocate to systemic tissues. Viable bacteria were found in spleen and were associated with increased abdominal fat pad mass in Zg16(-/-) animals. The function of ZG16 reveals a mechanism for keeping bacteria further away from the host colon epithelium.

Concepts: DNA, Archaea, Bacteria, Amino acid, Enzyme, Cell wall, Bacterial cell structure, Peptidoglycan


The bacterial flagellar motor consists of a rotor and a dozen stator units and regulates the number of active stator units around the rotor in response to changes in the environment. The MotPS complex is a Na(+)-type stator unit in the Bacillus subtilis flagellar motor and binds to the peptidoglycan layer through the peptidoglycan-binding (PGB) domain of MotS to act as the stator. The MotPS complex is activated in response to an increase in the Na(+) concentration in the environment, but the mechanism of this activation has remained unknown. We report that activation occurs by a Na(+)-induced folding and dimer formation of the PGB domain of MotS, as revealed in real-time imaging by high-speed atomic force microscopy. The MotPS complex showed two distinct ellipsoid domains connected by a flexible linker. A smaller domain, corresponding to the PGB domain, became structured and unstructured in the presence and absence of 150 mM NaCl, respectively. When the amino-terminal portion of the PGB domain adopted a partially stretched conformation in the presence of NaCl, the center-to-center distance between these two domains increased by up to 5 nm, allowing the PGB domain to reach and bind to the peptidoglycan layer. We propose that assembly of the MotPS complex into a motor proceeds by means of Na(+)-induced structural transitions of its PGB domain.

Concepts: Archaea, Bacteria, Flagellum, Bacillus, Peptidoglycan, Bacillus subtilis, Firmicutes, Electric motor


Phages play key roles in the pathogenicity and adaptation of the human pathogen Staphylococcus aureus. However, little is known about the molecular recognition events that mediate phage adsorption to the surface of S. aureus. The lysogenic siphophage ϕ11 infects S. aureus SA113. It was shown previously that ϕ11 requires α- or β-N-acetylglucosamine (GlcNAc) moieties on cell wall teichoic acid (WTA) for adsorption. Gp45 was identified as the receptor binding protein (RBP) involved in this process and GlcNAc residues on WTA were found to be the key component of the ϕ11 receptor. Here we report the crystal structure of the RBP of ϕ11, which assembles into a large, multidomain homotrimer. Each monomer contains a five-bladed propeller domain with a cavity that could accommodate a GlcNAc moiety. An electron microscopy reconstruction of the ϕ11 host adhesion component, the baseplate, reveals that six RBP trimers are assembled around the baseplate core. The Gp45 and baseplate structures provide insights into the overall organization and molecular recognition process of the phage ϕ11 tail. This assembly is conserved among most glycan-recognizing Siphoviridae, and the RBP orientation would allow host adhesion and infection without an activation step.

Concepts: DNA, Bacteria, Microbiology, Staphylococcus aureus, Infection, Bacteriophage, Cell wall, Peptidoglycan


Most bacteria contain a peptidoglycan (PG) cell wall, which is critical for maintenance of shape and important for cell division. In contrast, Planctomycetes have been proposed to produce a proteinaceous cell wall devoid of PG. The apparent absence of PG has been used as an argument for the putative planctomycetal ancestry of all bacterial lineages. Here we show, employing multiple bioinformatic methods, that planctomycetal genomes encode proteins required for PG synthesis. Furthermore, we biochemically demonstrate the presence of the sugar and the peptide components of PG in Planctomycetes. In addition, light and electron microscopic experiments reveal planctomycetal PG sacculi that are susceptible to lysozyme treatment. Finally, cryo-electron tomography demonstrates that Planctomycetes possess a typical PG cell wall and that their cellular architecture is thus more similar to that of other Gram-negative bacteria. Our findings shed new light on the cellular architecture and cell division of the maverick Planctomycetes.

Concepts: DNA, Protein, Cell, Archaea, Bacteria, Amino acid, Cell wall, Peptidoglycan


Clostridium difficile infection (CDI) is a challenging threat to human health. Infections occur after disruption of the normal microbiota, most commonly through the use of antibiotics. Current treatment for CDI largely relies on the broad-spectrum antibiotics vancomycin and metronidazole that further disrupt the microbiota resulting in frequent recurrence, highlighting the need for C. difficile-specific antimicrobials. The cell surface of C. difficile represents a promising target for the development of new drugs. C. difficile possesses a highly deacetylated peptidoglycan cell wall containing unique secondary cell wall polymers. Bound to the cell wall is an essential S-layer, formed of SlpA and decorated with an additional 28 related proteins. In addition to the S-layer, many other cell surface proteins have been identified, including several with roles in host colonization. This review aims to summarize our current understanding of these different C. difficile cell surface components and their viability as therapeutic targets.

Concepts: Medicine, Archaea, Bacteria, Cell membrane, Cell wall, Penicillin, Peptidoglycan, Clostridium difficile


Lysozyme is a cornerstone of innate immunity. The canonical mechanism for bacterial killing by lysozyme occurs through the hydrolysis of cell wall peptidoglycan (PG). Conventional type (c-type) lysozymes are also highly cationic and can kill certain bacteria independently of PG hydrolytic activity. Reflecting the ongoing arms race between host and invading microorganisms, both gram-positive and gram-negative bacteria have evolved mechanisms to thwart killing by lysozyme. In addition to its direct antimicrobial role, more recent evidence has shown that lysozyme modulates the host immune response to infection. The degradation and lysis of bacteria by lysozyme enhance the release of bacterial products, including PG, that activate pattern recognition receptors in host cells. Yet paradoxically, lysozyme is important for the resolution of inflammation at mucosal sites. This review will highlight recent advances in our understanding of the diverse mechanisms that bacteria use to protect themselves against lysozyme, the intriguing immunomodulatory function of lysozyme, and the relationship between these features in the context of infection.

Concepts: Immune system, Archaea, Bacteria, Enzyme, Microbiology, Innate immune system, Cell wall, Peptidoglycan