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Concept: Bacterial cell structure


Crotalicidin (Ctn), a cathelicidin-related peptide from the venom of a South American rattlesnake, possesses potent antimicrobial, antitumor, and antifungal properties. Previously, we have shown that its C-terminal fragment, Ctn[15-34], retains the antimicrobial and antitumor activities, but is less toxic to healthy cells and has improved serum stability. Here, we investigated the mechanism of action of Ctn and Ctn[15-34] against gram-negative bacteria. Both peptides were bactericidal, killing ~90% of Escherichia coli and Pseudomonas aeruginosa cells within 90-120 min and 5-30 min, respectively. Studies of ζ potential at the bacterial cell membrane suggested that both peptides accumulate at and neutralize negative charges on the bacterial surface. Flow cytometry experiments confirmed that both peptides permeabilize the bacterial cell membrane, but suggested slightly different mechanisms of action. Ctn[15-34] permeabilized the membrane immediately upon addition to the cells, whereas Ctn had a lag phase before inducing membrane damage and exhibited more complex cell-killing activity, probably because of two different modes of membrane permeabilization. Using surface plasmon resonance and leakage assays with model vesicles, we confirmed that Ctn[15-34] binds to and disrupts lipid membranes and also observed that Ctn[15-34] has a preference for vesicles that mimic bacterial or tumor cell membranes. Atomic force microscopy visualized the effect of these peptides on bacterial cells, and confocal microscopy confirmed their localization on the bacterial surface. Our studies shed light onto the antimicrobial mechanisms of Ctn and Ctn[15-34], suggesting Ctn[15-34] as a promising lead for development as an antibacterial/antitumor agent.

Concepts: Immune system, Protein, Bacteria, Cell membrane, Escherichia coli, Pseudomonas aeruginosa, Cell wall, Bacterial cell structure


The cell envelope of gram-negative bacteria, a structure comprising an outer (OM) and an inner (IM) membrane, is essential for life. The OM and the IM are separated by the periplasm, a compartment that contains the peptidoglycan. The OM is tethered to the peptidoglycan via the lipoprotein, Lpp. However, the importance of the envelope’s multilayered architecture remains unknown. Here, when we removed physical coupling between the OM and the peptidoglycan, cells lost the ability to sense defects in envelope integrity. Further experiments revealed that the critical parameter for the transmission of stress signals from the envelope to the cytoplasm, where cellular behaviour is controlled, is the IM-to-OM distance. Augmenting this distance by increasing the length of the lipoprotein Lpp destroyed signalling, whereas simultaneously increasing the length of the stress-sensing lipoprotein RcsF restored signalling. Our results demonstrate the physiological importance of the size of the periplasm. They also reveal that strict control over the IM-to-OM distance is required for effective envelope surveillance and protection, suggesting that cellular architecture and the structure of transenvelope protein complexes have been evolutionarily co-optimised for correct function. Similar strategies are likely at play in cellular compartments surrounded by 2 concentric membranes, such as chloroplasts and mitochondria.

Concepts: DNA, Cell, Bacteria, Eukaryote, Cell membrane, Cytoplasm, Cell wall, Bacterial cell structure


Biopolymer composite cell walls maintain cell shape and resist forces in plants, fungi and bacteria. Peptidoglycan, a crucial antibiotic target and immunomodulator, performs this role in bacteria. The textbook structural model of peptidoglycan is a highly ordered, crystalline material. Here we use atomic force microscopy (AFM) to image individual glycan chains in peptidoglycan from Escherichia coli in unprecedented detail. We quantify and map the extent to which chains are oriented in a similar direction (orientational order), showing it is much less ordered than previously depicted. Combining AFM with size exclusion chromatography, we reveal glycan chains up to 200 nm long. We show that altered cell shape is associated with substantial changes in peptidoglycan biophysical properties. Glycans from E. coli in its normal rod shape are long and circumferentially oriented, but when a spheroid shape is induced (chemically or genetically) glycans become short and disordered.

Concepts: Protein, Genetics, Archaea, Bacteria, Molecular biology, Escherichia coli, Cell wall, Bacterial cell structure


D-cycloserine is an antibiotic which targets sequential bacterial cell wall peptidoglycan biosynthesis enzymes: alanine racemase and D-alanine:D-alanine ligase. By a combination of structural, chemical and mechanistic studies here we show that the inhibition of D-alanine:D-alanine ligase by the antibiotic D-cycloserine proceeds via a distinct phosphorylated form of the drug. This mechanistic insight reveals a bimodal mechanism of action for a single antibiotic on different enzyme targets and has significance for the design of future inhibitor molecules based on this chemical structure.

Concepts: Cell, Bacteria, Amino acid, Adenosine triphosphate, Enzyme, Cell wall, Bacterial cell structure, Penicillin


Daptomycin is a highly efficient last-resort antibiotic that targets the bacterial cell membrane. Despite its clinical importance, the exact mechanism by which daptomycin kills bacteria is not fully understood. Different experiments have led to different models, including (i) blockage of cell wall synthesis, (ii) membrane pore formation, and (iii) the generation of altered membrane curvature leading to aberrant recruitment of proteins. To determine which model is correct, we carried out a comprehensive mode-of-action study using the model organism Bacillus subtilis and different assays, including proteomics, ionomics, and fluorescence light microscopy. We found that daptomycin causes a gradual decrease in membrane potential but does not form discrete membrane pores. Although we found no evidence for altered membrane curvature, we confirmed that daptomycin inhibits cell wall synthesis. Interestingly, using different fluorescent lipid probes, we showed that binding of daptomycin led to a drastic rearrangement of fluid lipid domains, affecting overall membrane fluidity. Importantly, these changes resulted in the rapid detachment of the membrane-associated lipid II synthase MurG and the phospholipid synthase PlsX. Both proteins preferentially colocalize with fluid membrane microdomains. Delocalization of these proteins presumably is a key reason why daptomycin blocks cell wall synthesis. Finally, clustering of fluid lipids by daptomycin likely causes hydrophobic mismatches between fluid and more rigid membrane areas. This mismatch can facilitate proton leakage and may explain the gradual membrane depolarization observed with daptomycin. Targeting of fluid lipid domains has not been described before for antibiotics and adds another dimension to our understanding of membrane-active antibiotics.

Concepts: Protein, Cell, Bacteria, Cell membrane, Cell wall, Membrane biology, Bacterial cell structure, Lipid bilayer


Bending over backward: Despite their small size, bacteria display highly organized cytoskeletal structures. Using microfabricated supports for model membranes, mechanical features of FtsZ (blue hexagons) filaments, a key component of bacterial cell division, can be addressed. Studying the curvature of an FtsZ filament into a groove or around a capillary helps to understand its mechanics.

Concepts: Cell, Bacteria, Eukaryote, Cell membrane, Cytoskeleton, Bacterial cell structure, Binary fission, FtsZ


Ultrasound coupling gel may serve as a vector for the spread of bacteria and has been the causative agent for significant health care-associated infections. The purpose of this study was to document existing infection-control procedures and level of contamination present within nonsterile ultrasound gel from several clinical departments at a single institution. A second purpose was to examine the effectiveness of clinician education and manufacturer-based ultrasound additives on ultrasound gel contamination and in vitro bacterial proliferation, respectively.

Concepts: Archaea, Clinical trial, Bacteria, Microbiology, Virus, Prokaryote, Bacterial cell structure, Microorganism


New switchable hydrogels are developed. Under acidic conditions, hydrogels undergo self- cyclization and can catch and kill bacteria. Under neutral/basic conditions, hydrogels undergo ring-opening and can release killed bacterial cells and resist protein adsorption and bacterial attachment. Smart hydrogels also show a dramatically improved mechanical property which is highly desired for biomedical applications.

Concepts: DNA, Gene, Cell nucleus, Bacteria, Amino acid, Enzyme, Cell wall, Bacterial cell structure


AIM: Taking into account that a novel strain of Bacillus megaterium was isolated from Uyuni salt lake (Bolivia) in a previous work, the objectives of this new study were to determine the maximal PHB production potential of B. megaterium strain uyuni S29 in an industrial conventional media, the possibility that the strain accumulates different types of PHA, the cellular morphology during the biosynthesis process, and the characterization of the produced biopolymers. METHODS AND RESULTS: The microorganism was first tested in a 3L-bioreactor obtaining a high specific growth rate of 1.64 h(-1) . A second fed-batch experiment was carried out in shaking flasks, reaching up to 70% PHB of cell dry mass. The biosynthesised polymers were extracted by two different extraction procedures and characterized. The results showed that all of them were PHB with thermal properties different to the conventional PHB. The micrographs taken by TEM show the cell different cell morphology during the fermentation process. CONCLUSIONS: In this previous study, the strain not only grew properly in the industrial conditions proposed without spore formation, but it also produced and accumulated a large content of PHB, never reached before for its genus. Therefore, if the culture conditions can be optimised, the biopolymer production could be increased. SIGNIFICANCE AND IMPACT OF STUDY: The impact of the study have related to the area of the biomaterials and their production. The study provides new data related to the high production of PHB from the wild novel strain B. megaterium uyuni S29, the highest polymer accumulation for the genus Bacillus without spores formation. © 2013 The Authors Journal of Applied Microbiology © 2013 The Society for Applied Microbiology.

Concepts: Archaea, Bacteria, Microbiology, Polymer, Bacillus, Bacterial cell structure, Biopolymer, Bacillus megaterium


The bacterial cell surface plays a major role in the bacterial aggregation that in turn plays a positive role in affecting the bacterial dispersion and survival in soil and their ability to adhere to plant surfaces. Plant growth-promoting Methylobacterium strains, Methylobacterium goesingense CBMB5, Methylobacterium sp. CBMB12, Methylobacterium oryzae CBMB20, Methylobacterium fujisawaense CBMB37, M. oryzae CBMB110 and Methylobacterium suomiense CBMB120 were evaluated for aggregation efficiency. Aggregation occurred in all test strains under high C/N growth conditions, and the strain CBMB12 showed the highest aggregation of 53.4 % at 72 h. Disaggregation compound treatment studies revealed the role of protein-protein interaction in Methylobacterium strains except CBMB110 and CBMB120 strains, where a possible carbohydrate-protein interaction is suspected. Surface layer protein extraction by LiCl followed by SDS-PAGE analysis showed the presence of proteins at molecular weights ranging from 41 to 49 kDa. Methylobacterium strains under aggregated conditions showed increased hydrophobicity compared to the cells under standard grown conditions. A relatively higher hydrophobicity of 50.1 % as evident by the adhesion with xylene was observed with strain CBMB12 under aggregated condition. This study reports the aggregation ability in plant growth-promoting Methylobacterium strains and the possible involvement of cellular components and hydrophobicity in this phenomenon.

Concepts: DNA, Protein, Cell nucleus, Archaea, Bacteria, Molecular biology, Eukaryote, Bacterial cell structure