scholarly journals Granular Layer in the Periplasmic Space of Gram-Positive Bacteria and Fine Structures of Enterococcus gallinarum and Streptococcus gordonii Septa Revealed by Cryo-Electron Microscopy of Vitreous Sections

2006 ◽  
Vol 188 (18) ◽  
pp. 6652-6660 ◽  
Author(s):  
Benoît Zuber ◽  
Marisa Haenni ◽  
Tânia Ribeiro ◽  
Kathrin Minnig ◽  
Fátima Lopes ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Granular Layer ◽  
Streptococcus Gordonii ◽  
Periplasmic Space ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Cryo Electron Microscopy ◽  
Enterococcus Gallinarum ◽  
Peptidoglycan Layer

ABSTRACT High-resolution structural information on optimally preserved bacterial cells can be obtained with cryo-electron microscopy of vitreous sections. With the help of this technique, the existence of a periplasmic space between the plasma membrane and the thick peptidoglycan layer of the gram-positive bacteria Bacillus subtilis and Staphylococcus aureus was recently shown. This raises questions about the mode of polymerization of peptidoglycan. In the present study, we report the structure of the cell envelope of three gram-positive bacteria (B. subtilis, Streptococcus gordonii, and Enterococcus gallinarum). In the three cases, a previously undescribed granular layer adjacent to the plasma membrane is found in the periplasmic space. In order to better understand how nascent peptidoglycan is incorporated into the mature peptidoglycan, we investigated cellular regions known to represent the sites of cell wall production. Each of these sites possesses a specific structure. We propose a hypothetic model of peptidoglycan polymerization that accommodates these differences: peptidoglycan precursors could be exported from the cytoplasm to the periplasmic space, where they could diffuse until they would interact with the interface between the granular layer and the thick peptidoglycan layer. They could then polymerize with mature peptidoglycan. We report cytoplasmic structures at the E. gallinarum septum that could be interpreted as cytoskeletal elements driving cell division (FtsZ ring). Although immunoelectron microscopy and fluorescence microscopy studies have demonstrated the septal and cytoplasmic localization of FtsZ, direct visualization of in situ FtsZ filaments has not been obtained in any electron microscopy study of fixed and dehydrated bacteria.

Microanatomy of the Periplasm of Bacillus Licheniformis

1971 ◽  
Vol 29 ◽  
pp. 262-263
Author(s):  
B.K. Ghosh
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Bacillus Licheniformis ◽  
External Environment ◽  
Permeability Barrier ◽  
Periplasmic Space ◽  
Modified Technique ◽  
Gram Positive ◽  
Gram Negative ◽  
Gram Positive Bacteria

Periplasm of bacteria is the space outside the permeability barrier of plasma membrane but enclosed by the cell wall. The contents of this special milieu exterior could be regulated by the plasma membrane from the internal, and by the cell wall from the external environment of the cell. Unlike the gram-negative organism, the presence of this space in gram-positive bacteria is still controversial because it cannot be clearly demonstrated. We have shown the importance of some periplasmic bodies in the secretion of penicillinase from Bacillus licheniformis.In negatively stained specimens prepared by a modified technique (Figs. 1 and 2), periplasmic space (PS) contained two kinds of structures: (i) fibrils (F, 100 Å) running perpendicular to the cell wall from the protoplast and (ii) an array of vesicles of various sizes (V), which seem to have evaginated from the protoplast.

scholarly journals Lipoteichoic Acid Is a Major Component of the Bacillus subtilis Periplasm

2008 ◽  
Vol 190 (22) ◽  
pp. 7414-7418 ◽  
Author(s):  
Valério R. F. Matias ◽  
Terry J. Beveridge
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Lipoteichoic Acid ◽  
Periplasmic Space ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Cryo Electron Microscopy ◽  
Resolution Observation ◽  
Positively Charged Gold Nanoparticles ◽  
Positively Charged

ABSTRACT Cryo-electron microscopy (cryo-EM) of frozen-hydrated specimens allows high-resolution observation of structures in optimally preserved samples. In gram-positive bacteria, this method reveals the presence of a periplasmic space between the plasma membrane and an often differentiated cell wall matrix. Since virtually nothing is known about the composition of its constituent matter (i.e., the periplasm), it is still unclear what structures (or mechanism) sustain a gram-positive periplasmic space. Here we have used cryo-EM of frozen-hydrated sections in combination with various labels to probe the model gram-positive organism Bacillus subtilis for major periplasmic components. Incubation of cells with positively charged gold nanoparticles showed almost similar levels of gold binding to the periplasm and the cell wall. On cells whose cell walls were enzymatically hydrolyzed (i.e., on protoplasts), a surface diffuse layer extending ∼30 nm from the membrane was revealed. The thickness and density of this layer were not significantly altered after treatment with a nonspecific protease, whereas it was labeled with anti-lipoteichoic acid (LTA) antibodies conjugated to nanogold. Further, the LTA layer spans most of the thickness of the periplasmic space, which strongly suggests that LTA is a major component of the B. subtilis periplasm.

scholarly journals A CYTOCHEMICAL LOCALIZATION OF REDUCTIVE SITES IN A GRAM-POSITIVE BACTERIUM

1964 ◽  
Vol 20 (3) ◽  
pp. 361-375 ◽  
Author(s):  
Woutera van Iterson ◽  
W. Leene
Keyword(s):  
Plasma Membrane ◽  
Cell Periphery ◽  
Gram Positive ◽  
Cytochemical Localization ◽  
Potassium Tellurite ◽  
Thin Sections ◽  
Gram Positive Bacteria ◽  
Cytoplasmic Membranes ◽  
Reducing Activity ◽  
The One

In bacteria the exact location of a respiratory enzyme system comparable to that of the mitochondria of other cells has remained uncertain. On the one hand, the existence of particulate "bacterial mitochondria" has been advocated (Mudd); on the other hand, important enzymes of the respiratory chain were recovered in the cytoplasmic membranes associated with some granular material (Weibull). In order to gain insight into this question, sites of reducing activity were localized in thin sections of bacteria using the reduction of potassium tellurite as an indicator. When this salt was added to the culture medium of Bacillus subtilis, it turned out that in this Gram-positive organism the reduced product is strictly bound at two sites, and that the plasma membrane does not materially gain in electron opacity through deposition of the reduced product. The reduction product is found on or in the membranes of particular organelles, which may possibly be regarded as the mitochondrial equivalents in Gram-positive bacteria, and which are sometimes seen connected to the plasma membrane. The second location is in thin rod-like elements at the cell periphery, possibly the sites from which the flagella emerge.

scholarly journals How Teichoic Acids Could Support a Periplasm in Gram-Positive Bacteria, and Let Cell Division Cheat Turgor Pressure

2021 ◽  
Vol 12 ◽  
Author(s):  
Harold P. Erickson
Keyword(s):  
Cell Division ◽  
Charge Density ◽  
Turgor Pressure ◽  
Support Pressure ◽  
Periplasmic Space ◽  
Gram Positive ◽  
Teichoic Acids ◽  
Gram Positive Bacteria ◽  
Peptidoglycan Cell Wall ◽  
Anionic Charge

The cytoplasm of bacteria is maintained at a higher osmolality than the growth medium, which generates a turgor pressure. The cell membrane (CM) cannot support a large turgor, so there are two possibilities for transferring the pressure to the peptidoglycan cell wall (PGW): (1) the CM could be pressed directly against the PGW, or (2) the CM could be separated from the PGW by a periplasmic space that is isoosmotic with the cytoplasm. There is strong evidence for gram-negative bacteria that a periplasm exists and is isoosmotic with the cytoplasm. No comparable studies have been done for gram-positive bacteria. Here I suggest that a periplasmic space is probably essential in order for the periplasmic proteins to function, including especially the PBPs that remodel the peptidoglycan wall. I then present a semi-quantitative analysis of how teichoic acids could support a periplasm that is isoosmotic with the cytoplasm. The fixed anionic charge density of teichoic acids in the periplasm is ∼0.5 M, which would bring in ∼0.5 M Na+ neutralizing ions. This approximately balances the excess osmolality of the cytoplasm that would produce a turgor pressure of 19 atm. The 0.5 M fixed charge density is similar to that of proteoglycans in articular cartilage, suggesting a comparability ability to support pressure. An isoosmotic periplasm would be especially important for cell division, since it would allow CM constriction and PGW synthesis to avoid turgor pressure.

scholarly journals Architecture of the human PI4KIIIα lipid kinase complex

2017 ◽  
Vol 114 (52) ◽  
pp. 13720-13725 ◽  
Author(s):  
Joshua A. Lees ◽  
Yixiao Zhang ◽  
Michael S. Oh ◽  
Curtis M. Schauder ◽  
Xiaoling Yu ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Active Sites ◽  
Structural Complexity ◽  
Signaling Molecules ◽  
Cell Physiology ◽  
Lipid Kinase ◽  
Membrane Interaction ◽  
Regulatory Subunits ◽  
Cryo Electron Microscopy

Plasma membrane (PM) phosphoinositides play essential roles in cell physiology, serving as both markers of membrane identity and signaling molecules central to the cell’s interaction with its environment. The first step in PM phosphoinositide synthesis is the conversion of phosphatidylinositol (PI) to PI4P, the precursor of PI(4,5)P2 and PI(3,4,5)P3. This conversion is catalyzed by the PI4KIIIα complex, comprising a lipid kinase, PI4KIIIα, and two regulatory subunits, TTC7 and FAM126. We here report the structure of this complex at 3.6-Å resolution, determined by cryo-electron microscopy. The proteins form an obligate ∼700-kDa superassembly with a broad surface suitable for membrane interaction, toward which the kinase active sites are oriented. The structural complexity of the assembly highlights PI4P synthesis as a major regulatory junction in PM phosphoinositide homeostasis. Our studies provide a framework for further exploring the mechanisms underlying PM phosphoinositide regulation.

Bactericidal effect of gentamicin-induced membrane vesicles derived fromPseudomonas aeruginosaPAO1 on gram-positive bacteria

2002 ◽  
Vol 48 (9) ◽  
pp. 810-820 ◽  
Author(s):  
Kelly L MacDonald ◽  
Terry J Beveridge
Keyword(s):  
Electron Microscopy ◽  
Therapeutic Potential ◽  
Hydrophobic Interaction Chromatography ◽  
Membrane Vesicles ◽  
Bactericidal Effect ◽  
Gram Positive ◽  
Thin Sections ◽  
Gram Negative ◽  
Induced Membrane ◽  
Gram Positive Bacteria

Previous studies have shown that gentamicin-induced membrane vesicles (g-MVs) from Pseudomonas aeruginosa PAO1 possess both the antibiotic (gentamicin) and a potent peptidoglycan hydrolase (PGase; autolysin) that is effective in killing gram-negative pathogens. This present study evaluated the therapeutic potential of g-MVs against four gram-positive bacteria. Bactericidal assays and electron microscopy of thin sections revealed that Bacillus subtilis 168 and Staphylococcus aureus D2C were susceptible to killing mediated by g-MVs, Listeria monocytogenes ATCC 19113 was slightly susceptible, whereas Enterococcus hirae ATCC 9790 was unaffected. g-MVs were generally more effective against the bacteria than was soluble gentamicin, suggesting they could have more killing power than natural membrane vesicles containing no antibiotic. Electron microscopy and hydrophobic interaction chromatography showed that more membrane vesicles (MVs) initially attached to B. subtilis (hydrophilic) than to predominantly hydrophobic E. hirae, L. monocytogenes, and S. aureus. Zymograms containing murein sacculi as an enzyme substrate illustrated that all organisms except E. hirae were sensitive to the 26-kDa autolysin to varying degrees. Peptidoglycan O-acetylation did not influence susceptibility to MV-mediated lysis. Though not universally effective, the g-MV delivery system remains a promising therapeutic alternative for specific gram-positive infections.Key words: gram-negative membrane vesicles, gentamicin, autolysin.

scholarly journals Structural basis of trehalose recycling by the ABC transporter LpqY-SugABC

2020 ◽  
Vol 6 (44) ◽  
pp. eabb9833
Author(s):  
Fengjiang Liu ◽  
Jingxi Liang ◽  
Bing Zhang ◽  
Yan Gao ◽  
Xiuna Yang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Abc Transporter ◽  
Abc Transporters ◽  
Unique Feature ◽  
Structural Basis ◽  
Human Pathogens ◽  
Gram Positive Bacteria ◽  
Cryo Electron Microscopy ◽  
Uptake Of Nutrients ◽  
Initial Mode

In bacteria, adenosine 5′-triphosphate (ATP)–binding cassette (ABC) importers are essential for the uptake of nutrients including the nonreducing disaccharide trehalose, a metabolite that is crucial for the survival and virulence of several human pathogens including Mycobacterium tuberculosis. SugABC is an ABC transporter that translocates trehalose from the periplasmic lipoprotein LpqY into the cytoplasm of mycobacteria. Here, we report four high-resolution cryo–electron microscopy structures of the mycobacterial LpqY-SugABC complex to reveal how it binds and passes trehalose through the membrane to the cytoplasm. A unique feature observed in this system is the initial mode of capture of the trehalose at the LpqY interface. Uptake is achieved by a pivotal rotation of LpqY relative to SugABC, moving from an open and accessible conformation to a clamped conformation upon trehalose binding. These findings enrich our understanding as to how ABC transporters facilitate substrate transport across the membrane in Gram-positive bacteria.

scholarly journals Architecture of the AP2:clathrin coat on the membranes of clathrin-coated vesicles

2020 ◽  
Author(s):  
Oleksiy Kovtun ◽  
Veronica Kane Dickson ◽  
Bernard T. Kelly ◽  
David. J. Owen ◽  
John A. G. Briggs
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Protein Composition ◽  
Binding Mode ◽  
Coated Vesicles ◽  
Conformational Rearrangement ◽  
Multiple Interfaces ◽  
Cryo Electron Microscopy ◽  
Subtomogram Averaging ◽  
Key Steps

AbstractClathrin-mediated endocytosis (CME) is crucial for modulating the protein composition of a cell’s plasma membrane. Clathrin forms a cage-like, polyhedral outer scaffold around a vesicle, to which cargo-selecting clathrin adaptors are attached. AP2 is the key adaptor in CME. Crystallography has shown AP2 to adopt a range of conformations. Here we used cryo-electron microscopy, tomography and subtomogram averaging to determine structures, interactions and arrangements of clathrin and AP2 at the key steps of coat assembly, from AP2 in solution to membrane-assembled clathrin-coated vesicles (CCVs). AP2 binds cargo and PtdIns(4,5)P2-containing membranes via multiple interfaces, undergoing conformational rearrangement from its cytosolic state. The binding mode of AP2 β2-appendage into the clathrin lattice in CCVs and buds implies how the adaptor structurally modulates coat curvature and coat disassembly.

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