How might a microbial cell that is entirely metabolically dormant – and which has the ability to remain so for extended periods of time – irreversibly commit itself to resuming vegetative growth ...within seconds of being exposed to certain amino acids or sugars? That this process takes place in the absence of any detectable ATP or de novo protein synthesis, and relies upon a pre-formed apparatus that is immobilised, respectively, in a semi-crystalline membrane or multi-layered proteinaceous coat, only exacerbates the challenge facing spores of Bacillales species when stimulated to germinate. Whereas the process by which spores are formed in response to nutrient starvation – sporulation – involves the orchestrated interplay between hundreds of distinct proteins, the process by which spores return to life – germination – is a much simpler affair, requiring a handful of receptor and channel proteins complemented with specialized peptidoglycan lysins. Despite this relative simplicity, and research effort spanning many decades, comprehensive understanding of key molecular and biochemical details and, in particular signal transduction mechanisms associated with spore germination, has remained elusive. In this review we provide an up to date overview of the field while identifying what we consider to be the key gaps in knowledge associated with germination of Bacillales spores, suggesting also technical approaches that may provide fresh insight to this unique biological process.
Spores of many species of the orders Bacillales and Clostridiales can be vectors for food spoilage, human diseases and intoxications, and biological warfare. Many agents are used for spore killing, ...including moist heat in an autoclave, dry heat at elevated temperatures, UV radiation at 254 and more recently 222 and 400 nm, ionizing radiation of various types, high hydrostatic pressures and a host of chemical decontaminants. An alternative strategy is to trigger spore germination, as germinated spores are much easier to kill than the highly resistant dormant spores—the so called “germinate to eradicate” strategy. Factors important to consider in choosing methods for spore killing include the: (1) cost; (2) killing efficacy and kinetics; (3) ability to decontaminate large areas in buildings or outside; and (4) compatibility of killing regimens with the: (i) presence of people; (ii) food quality; (iii) presence of significant amounts of organic matter; and (iv) minimal damage to equipment in the decontamination zone. This review will summarize research on spore killing and point out some common flaws which can make results from spore killing research questionable.
Piezoelectric materials are widely referred to as “smart” materials because they can transduce mechanical pressure acting on them to electrical signals and vice versa. They are extensively utilized ...in harvesting mechanical energy from vibrations, human motion, mechanical loads, etc., and converting them into electrical energy for low power devices. Piezoelectric transduction offers high scalability, simple device designs, and high‐power densities compared to electro‐magnetic/static and triboelectric transducers. This review aims to give a holistic overview of recent developments in piezoelectric nanostructured materials, polymers, polymer nanocomposites, and piezoelectric films for implementation in energy harvesting. The progress in fabrication techniques, morphology, piezoelectric properties, energy harvesting performance, and underpinning fundamental mechanisms for each class of materials, including polymer nanocomposites using conducting, non‐conducting, and hybrid fillers are discussed. The emergent application horizon of piezoelectric energy harvesters particularly for wireless devices and self‐powered sensors is highlighted, and the current challenges and future prospects are critically discussed.
This paper presents a comprehensive review of the energy harvesting performance of different types of piezoelectric materials. These materials include nanostructured materials, polymers, polymer nanocomposites synthesized using different types of fillers and piezoelectric films. The fabrication techniques, energy harvesting mechanisms, and applications of piezoelectric nanogenerators built using these materials are discussed thoroughly.
Bacteria belonging to the orders Bacillales and Clostridiales form spores in response to nutrient starvation. From a simplified morphological perspective, the spore can be considered as comprising a ...central protoplast or core, that is, enveloped sequentially by an inner membrane (IM), a peptidoglycan cortex, an outer membrane, and a proteinaceous coat. All of these structures are characterized by unique morphological and/or structural features, which collectively confer metabolic dormancy and properties of environmental resistance to the quiescent spore. These properties are maintained until the spore is stimulated to germinate, outgrow and form a new vegetative cell. Spore germination comprises a series of partially overlapping biochemical and biophysical events – efflux of ions from the core, rehydration and IM reorganization, disassembly of cortex and coat – all of which appear to take place in the absence of
de novo
ATP and protein synthesis. If the latter points are correct, why then do spores of all species examined to date contain a diverse range of mRNA molecules deposited within the spore core? Are some of these molecules “functional,” serving as translationally active units that are required for efficient spore germination and outgrowth, or are they just remnants from sporulation whose sole purpose is to provide a reservoir of ribonucleotides for the newly outgrowing cell? What is the fate of these molecules during spore senescence, and indeed, are conditions within the spore core likely to provide any opportunity for changes in the transcriptional profile of the spore during dormancy? This review encompasses a historical perspective of spore ribonucleotide biology, from the earliest biochemical led analyses – some of which in hindsight have proved to be remarkably prescient – through the transcriptomic era at the turn of this century, to the latest next generation sequencing derived insights. We provide an overview of the key literature to facilitate reasoned responses to the aforementioned questions, and many others, prior to concluding by identifying the major outstanding issues in this crucial area of spore biology.
Recombinant enzyme expression in Escherichia coli is one of the most popular methods to produce bulk concentrations of protein product. However, this method is often limited by the inadvertent ...formation of inclusion bodies. Our analysis systematically reviews literature from 2010 to 2021 and details the methods and strategies researchers have utilized for expression of difficult to express (DtE), industrially relevant recombinant enzymes in E. coli expression strains. Our review identifies an absence of a coherent strategy with disparate practices being used to promote solubility. We discuss the potential to approach recombinant expression systematically, with the aid of modern bioinformatics, modelling, and 'omics' based systems-level analysis techniques to provide a structured, holistic approach. Our analysis also identifies potential gaps in the methods used to report metadata in publications and the impact on the reproducibility and growth of the research in this field.
We report the use of DNA origami nanostructures, functionalized with aptamers, as a vehicle for delivering the antibacterial enzyme lysozyme in a specific and efficient manner. We test the system ...against Gram‐positive (Bacillus subtilis) and Gram‐negative (Escherichia coli) targets. We use direct stochastic optical reconstruction microscopy (dSTORM) and atomic force microscopy (AFM) to characterize the DNA origami nanostructures and structured illumination microscopy (SIM) to assess the binding of the origami to the bacteria. We show that treatment with lysozyme‐functionalized origami slows bacterial growth more effectively than treatment with free lysozyme. Our study introduces DNA origami as a tool in the fight against antibiotic resistance, and our results demonstrate the specificity and efficiency of the nanostructure as a drug delivery vehicle.
Antibiotic resistance is a growing health issue that is now rendering humans vulnerable once again to infections that have been treatable for decades. Various approaches have been proposed to overcome this threat and effectively treat bacterial infections. DNA nanostructures, functionalized with aptamers, were used as a vehicle for delivering the antibacterial enzyme lysozyme in a specific and efficient manner, to destroy bacterial targets.
Enzyme therapies are attracting significant attention as thrombolytic drugs during the current scenario owing to their great affinity, specificity, catalytic activity, and stability. Among various ...sources, the application of microbial-derived thrombolytic and fibrinolytic enzymes to prevent and treat vascular occlusion is promising due to their advantageous cost–benefit ratio and large-scale production. Thrombotic complications such as stroke, myocardial infarction, pulmonary embolism, deep venous thrombosis, and peripheral occlusive diseases resulting from blood vessel blockage are the major cause of poor prognosis and mortality. Given the ability of microbial thrombolytic enzymes to dissolve blood clots and prevent any adverse effects, their use as a potential thrombolytic therapy has attracted great interest. A better understanding of the hemostasis and fibrinolytic system may aid in improving the efficacy and safety of this treatment approach over classical thrombolytic agents. Here, we concisely discuss the physiological mechanism of thrombus formation, thrombo-, and fibrinolysis, thrombolytic and fibrinolytic agents isolated from bacteria, fungi, and algae along with their mode of action and the potential application of microbial enzymes in thrombosis therapy.
The quest for bacterial survival is exemplified by spores formed by some
members. They turn up everywhere one looks, and their ubiquity reflects adaptations to the stresses bacteria face. Spores are ...impactful in public health, food safety, and biowarfare. Heat resistance is the hallmark of spores and is countered principally by a mineralized gel-like protoplast, termed the spore core, with reduced water which minimizes macromolecular movement/denaturation/aggregation. Dry heat, however, introduces mutations into spore DNA. Spores have countermeasures to extreme conditions that are multifactorial, but the fact that spore DNA is in a crystalline-like nucleoid in the spore core, likely due to DNA saturation with small acid-soluble spore proteins (SASPs), suggests that reduced macromolecular motion is also critical in spore dry heat resistance. SASPs are also central in the radiation resistance characteristic of spores, where the contributions of four spore features-SASP; Ca
, with pyridine-2,6-dicarboxylic acid (CaDPA); photoproduct lyase; and low water content-minimize DNA damage. Notably, the spore environment steers UV photochemistry toward a product that germinated spores can repair without significant mutagenesis. This resistance extends to chemicals and macromolecules that could damage spores. Macromolecules are excluded by the spore coat which impedes the passage of moieties of ≥10 kDa. Additionally, damaging chemicals may be degraded or neutralized by coat enzymes/proteins. However, the principal protective mechanism here is the inner membrane, a compressed structure lacking lipid fluidity and presenting a barrier to the diffusion of chemicals into the spore core; SASP saturation of DNA also protects against genotoxic chemicals. Spores are also resistant to other stresses, including high pressure and abrasion. Regardless, overarching mechanisms associated with resistance seem to revolve around reduced molecular motion, a fine balance between rigidity and flexibility, and perhaps efficient repair.
The exosporium of
QM B1551 spores is morphologically distinct to exosporia observed in spores of many other species. Previous work has demonstrated that unidentified genes encoded on one of the large ...indigenous plasmids are required for the assembly of the
exosporium. Here we provide evidence that pBM600-encoded orthologues of the
CotW and CotX proteins, which form the crust layer in spores of that species, are structural components of the
QM B1551 spore exosporium. The introduction of plasmid-borne
and orthologous
genes to the PV361 strain, which lacks all indigenous plasmids and produces spores that are devoid of an exosporium, results in the development of spores with a rudimentary exosporium-type structure. Additionally, purified recombinant CotW protein is shown to assemble at the air-water interface to form thin sheets of material, which is consistent with the idea that this protein may form a basal layer in the
QM B1551 exosporium.
When starved of nutrients some bacterial species develop metabolically dormant spores that can persist in a viable state in the environment for several years. The outermost layers of spores are of particular interest since (a) these represent the primary site for interaction with the environment, and (b) the protein constituents may have biotechnological applications. The outermost layer, or exosporium, in
QM B1551 spores, is of interest as it is morphologically distinct to the exosporium of spores of the pathogenic
family. In this work we provide evidence that structurally important protein constituents of the
exosporium are different to those in the
family. We show also that one of these proteins can assemble when purified to form sheets of exosporium-like material. This is significant as it indicates that spore-forming bacteria employ different proteins, and mechanisms of assembly, to construct their external layers.
The world faces a major infectious disease challenge. Interest in the discovery, design, or development of antimicrobial peptides (AMPs) as an alternative approach for the treatment of bacterial ...infections has increased. Insects are a good source of AMPs which are the main effector molecules of their innate immune system. Black Soldier Fly Larvae (BSFL) are being developed for large-scale rearing for food sustainability, waste reduction and as sustainable animal and fish feed. Bioinformatic studies have suggested that BSFL have the largest number of AMPs identified in insects. However, most AMPs identified in BSF have not yet undergone antimicrobial evaluation but are promising leads to treat critical infections.
Jg7197.t1, Jg7902.t1 and Jg7904.t1 were expressed into the haemolymph of larvae following infection with Salmonella enterica serovar Typhimurium and were predicted to be AMPs using the computational tool ampir. The genes encoding these proteins were within 2 distinct clusters in chromosome 1 of the BSF genome. Following removal of signal peptides, predicted structures of the mature proteins were superimposed, highlighting a high degree of structural conservation. The 3 AMPs share primary sequences with proteins that contain a Kunitz-binding domain; characterised for inhibitory action against proteases, and antimicrobial activities. An in vitro antimicrobial screen indicated that heterologously expressed SUMO-Jg7197.t1 and SUMO-Jg7902.t1 did not show activity against 12 bacterial strains. While recombinant SUMO-Jg7904.t1 had antimicrobial activity against a range of Gram-negative and Gram-positive bacteria, including the serious pathogen Pseudomonas aeruginosa.
We have cloned and purified putative AMPs from BSFL and performed initial in vitro experiments to evaluate their antimicrobial activity. In doing so, we have identified a putative novel defensin-like AMP, Jg7904.t1, encoded in a paralogous gene cluster, with antimicrobial activity against P. aeruginosa.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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