Cell wall assembly in Bacillus subtilis: how spirals and spaces challenge paradigms (original) (raw)
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Journal of Bacteriology, 1991
The pattern of cross-linking in the peptidoglycan of Bacillus megaterium has been studied by the pulsed addition of radiolabeled diaminopimelic acid. The distribution of label in muropeptides, generated by digestion with Chalaropsis muramidase and separated by high-performance liquid chromatography, stabilized after 0.15 of a generation time. The proportion of label in the acceptor and donor positions of isolated muropeptide dimers stabilized over the same period of time. The results have led to the formulation a new model for the assembly of peptidoglycan into the cylindrical wall of B. megaterium by a monomer addition process. Single nascent glycan peptide strands form cross-linkages only with material at the inner surface of the wall. Maturation is a direct consequence of subsequent incorporation of further new glycan peptide strands, and there is no secondary cross-linking process. The initial distribution of muropeptides is constant. It follows that the final pattern of cross-l...
Cell wall synthesis is necessary for membrane dynamics during sporulation of Bacillus subtilis
Molecular Microbiology, 2010
During Bacillus subtilis sporulation, an endocytic-like process called engulfment results in one cell being entirely encased in the cytoplasm of another cell. The driving force underlying this process of membrane movement has remained unclear, although components of the machinery have been characterized. Here we provide evidence that synthesis of peptidoglycan, the rigid, strength bearing extracellular polymer of bacteria, is a key part of the missing force-generating mechanism for engulfment. We observed that sites of peptidoglycan synthesis initially coincide with the engulfing membrane and later with the site of engulfment membrane fission. Furthermore, compounds that block muropeptide synthesis or polymerization prevented membrane migration in cells lacking a component of the engulfment machinery (SpoIIQ), and blocked the membrane fission event at the completion of engulfment in all cells. In addition, these compounds inhibited bulge and vesicle formation that occur in spoIID mutant cells unable to initiate engulfment, as did genetic ablation of a protein that polymerizes muropeptides. This is the first report to our knowledge that peptidoglycan synthesis is necessary for membrane movements in bacterial cells and has implications for the mechanism of force generation during cytokinesis.
Microbiology, 1989
Uranyl actetate staining of thin sections allowed a distinction to be made between cell wall material that contains teichoic acid and that which contains teichuronic acid. The stain was used to study the pattern of wall assembly in Bacillus subtilis undergoing transitions between growth conditions leading to incorporation of the different anionic polymers. The results showed that new material is incorporated along the inner surface of the cylindrical region of the wall confirming, by a more direct method, results obtained earlier with teichoic acid specific phages. New material appears to be evenly distributed along the inner surface and no evidence was obtained for the presence of specific zones of incorporation. Abbreviations: TA, teichoic acid; TU, teichuronic acid; TA or TU walls, cell wall material containing predominantly TA or TU as the anionic polymer, respectively.
Medical Microbiology and Immunology, 1999
The classical concept of the architecture of microbial murein assumes cross-linked glycan chains to be arranged in horizontal layers outside of the plasma membrane. It necessitates elaborate hypotheses to explain processes such as the biosynthesis, growth and division of the bacterial cell wall and provides no explanation for transenvelope macromolecular transport. Moreover, this model is difficult to reconcile with a number of basic chemical and electron microscopical data. According to a fundamentally distinct concept which is presented here, glycan strands in the microbial wall run perpendicular to the plasma membrane, each strand being cross-linked by peptide bridges with four other strands. This arrangement allows the formation of a structured matrix pierced with ordered ionophoric channels potentially harboring either lipoprotein or teichoic (lipoteichoic) acid molecules in Gram-negative and Gram-positive bacteria, respectively. New wall structures are synthesized in toto emerging from the cytoplasmic membrane as a condensed gel-like network below the old wall without being covalently attached to it, expanding due to inherent elasticity as the old wall is lyzed. This model reflects published genetic and biochemical data and offers a simple explanation for peptidoglycan biogenesis. As the biosynthesis is terminated by enzymic cleavage of all glycan strands, murein is irreversibly released from the membrane. The murein detachment prepares the membrane for de novo assembly of both the new wall synthesis machinery and the multicomponent factory for protein, DNA and phospholipid transfer. Being assembled in parallel, both new murein and the traffic complexes grow from the membrane together. This concept eliminates the necessity for the traffic complexes to penetrate intact murein. In the process of simultaneous assembly, the expanding murein functions as a lifting platform driven by the force of turgor pressure, transporting macromolecules through the perisplasmic space.
A widespread family of bacterial cell wall assembly proteins
The EMBO Journal, 2011
Teichoic acids and acidic capsular polysaccharides are major anionic cell wall polymers (APs) in many bacteria, with various critical cell functions, including maintenance of cell shape and structural integrity, charge and cation homeostasis, and multiple aspects of pathogenesis. We have identified the widespread LytR-Cps2A-Psr (LCP) protein family, of previously unknown function, as novel enzymes required for AP synthesis. Structural and biochemical analysis of several LCP proteins suggest that they carry out the final step of transferring APs from their lipid-linked precursor to cell wall peptidoglycan (PG). In Bacillus subtilis, LCP proteins are found in association with the MreB cytoskeleton, suggesting that MreB proteins coordinate the insertion of the major polymers, PG and AP, into the cell wall.
Cell wall-polypeptide complexes in Bacillus subtilis
Carbohydrate Research, 1983
The cell surface of Bacillus subtilis contains several peptidoglycan-associated polypeptides. Cell walls were labeled with 1251 or %, 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 [3"S]-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.
Towards a comprehensive view of the bacterial cell wall
Trends in Microbiology, 2005
Direct in vivo visualization, in full atomic detail, of the microbial cell wall and its stress-bearing structural architecture remains one of the prime challenges in microbiology. In the meantime, molecular modeling can provide a framework for explaining and predicting mechanisms involved in morphogenesis, bacterial cell growth and cell division, during which the wall and its major structural component -murein -have to protect the cell from osmotic pressure and multiple tensile forces. Here, we illustrate why the scaffold concept of murein architecture provides a more comprehensive representation of bacterial cell wall physiology than previous models.
Distinct and essential morphogenic functions for wall- and lipo-teichoic acids in Bacillus subtilis
The EMBO Journal, 2009
Teichoic acids (TAs) are anionic polymers that constitute a major component of the cell wall in most Gram-positive bacteria. Despite decades of study, their function has remained unclear. TAs are covalently linked either to the cell wall peptidoglycan (wall TA (WTA)) or to the membrane (lipo-TA (LTA)). We have characterized the key enzyme of LTA synthesis in Bacillus subtilis, LTA synthase (LtaS). We show that LTA is needed for divalent cation homoeostasis and that its absence has severe effects on cell morphogenesis and cell division. Inactivation of both LTA and WTA is lethal and comparison of the individual mutants suggests that they have differentiated roles in elongation (WTA) and division (LTA). B. subtilis has four ltaS paralogues and we show how their roles are partially differentiated. Two paralogues have a redundant role in LTA synthesis during sporulation and their absence gives a novel absolute block in sporulation. The crystal structure of the extracytoplasmic part of LtaS, solved at 2.4-Å resolution, reveals a phosphorylated threonine residue, which provides clues about the catalytic mechanism and identifies the active site of the enzyme.
Shape determination in Bacillus subtilis
Current Opinion in Microbiology, 2007
The discovery of cytoskeletal elements in prokaryotes has dramatically changed the way we think about bacterial cell morphogenesis. The rod shape of Bacillus subtilis is maintained by the two major polymers (peptidoglycan and teichoic acids) of its thick cell wall and determined by the way these are inserted during growth. The current view is that the dynamic tubulin-like (FtsZ) and actin-like (MreB) cytoskeletons orchestrate, both in time and space, the assembly of macromolecular machineries that effect cell wall synthesis and hydrolysis during cell division and cell elongation, respectively.