High-resolution analysis of the peptidoglycan composition inStreptomyces coelicolor

Abstract

22The bacterial cell wall maintains cell shape and protects against bursting by the 23 turgor. A major constituent of the cell wall is peptidoglycan (PG), which is 24 continuously modified to allow cell growth and differentiation through the concerted 25 activity of biosynthetic and hydrolytic enzymes. Streptomycetes are Gram-positive 26 bacteria with a complex multicellular life style alternating between mycelial growth 27 and the formation of reproductive spores. This involves cell-wall remodeling at apical 28 sites of the hyphae during cell elongation and autolytic degradation of the vegetative 29 mycelium during the onset of development and antibiotic production. Here, we show 30 that there are distinct differences in the cross-linking and maturation of the PG 31 between exponentially growing vegetative hyphae and the aerial hyphae that 32 undergo sporulation. LC-MS/MS analysis identified over 80 different muropeptides, 33 revealing that major PG hydrolysis takes place over the course of mycelial growth. 34 Half of the dimers lack one of the disaccharide units in transition-phase cells, most 35 likely due to autolytic activity. De-acetylation of MurNAc to MurN was particularly 36 pronounced in spores, suggesting that MurN plays a role in spore development. 37 Taken together, our work highlights dynamic and growth phase-dependent 38 construction and remodeling of PG in Streptomyces. 39 40 IMPORTANCE 41 Streptomycetes are bacteria with a complex lifestyle, which are model organisms for 42 bacterial multicellularity. From a single spore a large multigenomic, multicellular 43 mycelium is formed, which differentiates to form spores. Programmed cell death is an 44 important event during the onset of morphological differentiation. In this work we 45 3 provide new insights into the changes in the peptidoglycan architecture over time, 46 highlighting changes over the course of development and between growing mycelia 47 56 Peptidoglycan (PG) is a major component of the bacterial cell wall. It forms a physical 57boundary that maintains cell shape, protects cellular integrity against the osmotic 58 pressure and acts as a scaffold for large protein assemblies and exopolymers (66). 59The cell wall is a highly dynamic macromolecule that is continuously constructed and 60 deconstructed to allow for cell growth and to meet environmental demands (27). PG 61 is built up of glycan strands of alternating N-acetylglucosamine (GlcNAc) and N-62 acetylmuramic acid (MurNAc) residues that are connected by short peptides to form 63 a mesh-like polymer. PG biosynthesis starts with the synthesis of PG precursors by 64 the Mur enzymes in the cytoplasm and cell membrane, resulting in lipid II precursor, 65 undecaprenylpyrophosphoryl-MurNAc(GlcNAc)-pentapeptide. Lipid II is transported 66 across the cell membrane by MurJ and/or FtsW/SEDS proteins and the PG is 67 polymerized and incorporated into the existing cell wall by the activities of 68 glycosyltransferases and transpeptidases (9, 30, 57). 69 The Gram-positive model bacterium Bacillus ...

Ultrastructure of Exospore Formation in Streptomyces Revealed by Cryo-Electron Tomography

“…In some instances, two distinct spore wall layers could be identified - a darker inner layer (∼20-nm thick) and a lighter, outer layer (∼30-nm thick). The spore wall overall has an increased percentage of 3–4 crosslinks and decreased 3–3 crosslinks compared to the PG of aerial hyphae ( van der Aart et al, 2018 ), however, whether there are structural differences between the inner and outer layers in the spore wall remains unknown.…”

Alterations in the Cell Wall of Rhodococcus biphenylivorans Under Norfloxacin Stress

“…The resulting pellets were washed with ddH 2 O and freeze-dried for infrared spectroscopy analysis. The peptidoglycan isolation from TG9 cells in the three different states was performed as described previously ( Kuhner et al, 2014 ; Van der Aart et al, 2018 ), with some modifications. A total of 100 mL of the exponentially growing, VBNC, and resuscitation cultures were collected by centrifugation at 10,000 g for 20 min.…”