scholarly journals Controlled Autolysis and Enzyme Release in a Recombinant Lactococcal Strain Expressing the Metalloendopeptidase Enterolysin A

2004 ◽  
Vol 70 (3) ◽  
pp. 1744-1748 ◽  
Author(s):  
Rita M. Hickey ◽  
R. Paul Ross ◽  
Colin Hill
Keyword(s):  
Large Scale ◽  
Production Systems ◽  
Expression System ◽  
Fold Increase ◽  
Lytic Enzyme ◽  
Enzyme Release ◽  
Gram Positive Bacteria ◽  
Wide Range ◽  
Potential Applications ◽  
A Cell

ABSTRACT This study concerns the exploitation of the lytic enzyme enterolysin A (EntL), produced by Enterococcus faecalis strain DPC5280, to elicit the controlled autolysis of starter lactococci. EntL, a cell wall metalloendopeptidase secreted by some E. faecalis strains, can kill a wide range of gram-positive bacteria, including lactococci. The controlled expression of entL, which encodes EntL, was achieved using a nisin-inducible expression system in a lactococcal host. Zymographic analysis of EntL activity demonstrated that active enzyme is produced by the recombinant lactococcal host. Indeed, expression of EntL resulted in almost complete autolysis of the host strain 2 h after induction with nisin. Model cheese experiments using a starter strain in addition to the inducible enterolysin-producing strain showed a 27-fold increase in activity with respect to the release of lactate dehydrogenase in the strain overexpressing EntL, demonstrating the potential of EntL production in large-scale cheese production systems. Indeed, the observation that a wide range of lactic bacteria are sensitive to EntL suggests that EntL-induced autolysis has potential applications with a variety of lactic acid bacteria and could be a basis for probiotic delivery systems.

scholarly journals Sporulation in solventogenic and acetogenic clostridia

2021 ◽  
Author(s):  
Mamou Diallo ◽  
Servé W. M. Kengen ◽  
Ana M. López-Contreras
Keyword(s):  
Current Knowledge ◽  
Carbon Sources ◽  
Cost Effective ◽  
Biotechnological Production ◽  
Key Points ◽  
Wide Range ◽  
Potential Applications ◽  
A Cell ◽  
Sporulation Initiation ◽  
Solventogenic Clostridia

AbstractThe Clostridium genus harbors compelling organisms for biotechnological production processes; while acetogenic clostridia can fix C1-compounds to produce acetate and ethanol, solventogenic clostridia can utilize a wide range of carbon sources to produce commercially valuable carboxylic acids, alcohols, and ketones by fermentation. Despite their potential, the conversion by these bacteria of carbohydrates or C1 compounds to alcohols is not cost-effective enough to result in economically viable processes. Engineering solventogenic clostridia by impairing sporulation is one of the investigated approaches to improve solvent productivity. Sporulation is a cell differentiation process triggered in bacteria in response to exposure to environmental stressors. The generated spores are metabolically inactive but resistant to harsh conditions (UV, chemicals, heat, oxygen). In Firmicutes, sporulation has been mainly studied in bacilli and pathogenic clostridia, and our knowledge of sporulation in solvent-producing or acetogenic clostridia is limited. Still, sporulation is an integral part of the cellular physiology of clostridia; thus, understanding the regulation of sporulation and its connection to solvent production may give clues to improve the performance of solventogenic clostridia. This review aims to provide an overview of the triggers, characteristics, and regulatory mechanism of sporulation in solventogenic clostridia. Those are further compared to the current knowledge on sporulation in the industrially relevant acetogenic clostridia. Finally, the potential applications of spores for process improvement are discussed.Key Points• The regulatory network governing sporulation initiation varies in solventogenic clostridia.• Media composition and cell density are the main triggers of sporulation.• Spores can be used to improve the fermentation process.

scholarly journals Bioprocesses with Reduced Ecological Footprint by Marine Debaryomyces hansenii Strain for Potential Applications in Circular Economy

2021 ◽  
Vol 7 (12) ◽  
pp. 1028
Author(s):  
Silvia Donzella ◽  
Claudia Capusoni ◽  
Luisa Pellegrino ◽  
Concetta Compagno
Keyword(s):  
Ecological Footprint ◽  
Debaryomyces Hansenii ◽  
High Growth ◽  
High Growth Rate ◽  
Food Industries ◽  
Starting Point ◽  
Wide Range ◽  
Potential Applications ◽  
A Cell ◽  
Marine Strain

The possibility to perform bioprocesses with reduced ecological footprint to produce natural compounds and catalyzers of industrial interest is pushing the research for salt tolerant microorganisms able to grow on seawater-based media and able to use a wide range of nutrients coming from waste. In this study we focused our attention on a Debaryomyces hansenii marine strain (Mo40). We optimized cultivation in a bioreactor at low pH on seawater-based media containing a mixture of sugars (glucose and xylose) and urea. Under these conditions the strain exhibited high growth rate and biomass yield. In addition, we characterized potential applications of this yeast biomass in food/feed industry. We show that Mo40 can produce a biomass containing 45% proteins and 20% lipids. This strain is also able to degrade phytic acid by a cell-bound phytase activity. These features represent an appealing starting point for obtaining D. hansenii biomass in a cheap and environmentally friendly way, and for potential use as an additive or to replace unsustainable ingredients in the feed or food industries, as this species is included in the QPS EFSA list (Quality Presumption as Safe—European Food Safety Authority).

scholarly journals Characterization of Leader Processing Shows That Partially Processed Mersacidin Is Activated by AprE After Export

2021 ◽  
Vol 12 ◽  
Author(s):  
Jakob H. Viel ◽  
Amanda Y. van Tilburg ◽  
Oscar P. Kuipers
Keyword(s):  
Antimicrobial Activity ◽  
Synthetic Biology ◽  
Heterologous Expression ◽  
Expression System ◽  
Heterologous Expression System ◽  
Proteolytic Activation ◽  
E Coli ◽  
Gram Positive Bacteria ◽  
Precursor Peptide ◽  
Potential Applications

The ribosomally synthesized and post-translationally modified peptide mersacidin is a class II lanthipeptide with good activity against Gram-positive bacteria. The intramolecular lanthionine rings, that give mersacidin its stability and antimicrobial activity, are specific structures with potential applications in synthetic biology. To add the mersacidin modification enzymes to the synthetic biology toolbox, a heterologous expression system for mersacidin in Escherichia coli has recently been developed. While this system was able to produce fully modified mersacidin precursor peptide that could be activated by Bacillus amyloliquefaciens supernatant and showed that mersacidin was activated in an additional proteolytic step after transportation out of the cell, it lacked a mechanism for clean and straightforward leader processing. Here, the protease responsible for activating mersacidin was identified and heterologously produced in E. coli, improving the previously reported heterologous expression system. By screening multiple proteases, the stringency of proteolytic activity directly next to a very small lanthionine ring is demonstrated, and the full two-step proteolytic activation of mersacidin was elucidated. Additionally, the effect of partial leader processing on diffusion and antimicrobial activity is assessed, shedding light on the function of two-step leader processing.

Blueprints for Tropical Dairy Farming

2017 ◽  
Author(s):  
John Moran ◽  
Philip Chamberlain
Keyword(s):  
Developing Countries ◽  
Best Management Practices ◽  
Large Scale ◽  
Management Practices ◽  
Production Systems ◽  
Dairy Farming ◽  
Dairy Herds ◽  
Dairy Production ◽  
Wide Range ◽  
The Tropics

Blueprints for Tropical Dairy Farming provides insight into the logistics, infrastructure and management required for the development of small and large dairy farms in tropical developing countries. Farmers will learn how to improve the welfare, milk quality and productivity of their dairy herds. This book complements author John Moran’s five previous books on the principles of tropical dairy farming. The manual covers a wide range of topics related to ensuring the sustainability of dairy production systems in tropical developing countries, such as South and East Asia, Africa and Central America. It also provides guidelines for the best management practices of large-scale, more intensive dairy systems. While smallholder farms are the major suppliers of milk in the tropics, many larger farms are becoming established throughout the tropics to satisfy the increasing demands for fresh milk. Blueprints for Tropical Dairy Farming will be a valuable resource for farmers and stockpeople who want to improve the productive performance of their dairy herds, farm advisers who can assist farmers to achieve this aim, educators who develop training programs for farmers or who train dairy advisers in the basics of dairy production technology, and other stakeholders in tropical dairy production, such as local agribusiness, policy makers and research scientists. National and international agencies will learn new insights into the required long-term logistics for regional dairy development, while potential investors will acquire knowledge into intensive tropical dairy farming.

Simultaneously improving the ductility and strength of aromatic thermoset films

2021 ◽  
pp. 095400832110171
Author(s):  
Cheng Wang ◽  
Long Fei Zhang ◽  
Wa Li ◽  
Li Rong Yang ◽  
Jia Jun Ma ◽  
...  
Keyword(s):  
Electrostatic Interactions ◽  
Large Scale ◽  
Noncovalent Interactions ◽  
Ion Pair ◽  
Special Group ◽  
Covalent Bonds ◽  
New Class ◽  
Wide Range ◽  
Potential Applications ◽  
Mechanical Strengths

Aromatic thermoset materials have shown great potential applications in various fields owing to their excellent mechanical strengths. However, their poor ductility is still hinders their large-scale applications. In this study, a new class of aromatic thermosets consisting of two types of crosslinks was successfully developed by incorporating the special group imidazole into a type of crosslinked thermoset. One crosslink is constituted of reversible multiple noncovalent interactions containing “face-face” π–π stacking, “point-point” hydrogen bonds, and ion-pair electrostatic interactions, whereas the other is composed of permanent covalent bonds. Most importantly, the synergetic interplay among these reversible multiple noncovalent interactions enables them to evade the restrictions from the aromatic polymer skeletons to proceed with their dynamic dissociating-rebuilding processes, which can timely and effectively dissipate the internal stress. Finally, owing to the coefficient of these two types of crosslinks, a significantly enhanced ductility was successfully obtained on these aromatic thermosets and their tensile strengths were also improved. Such thermosets having simultaneously enhanced strengths and ductility are predicted to be eventually used in a wide range of applications.

scholarly journals The complete genome sequence of the nitrile biocatalyst Rhodocccus rhodochrous ATCC BAA-870

2019 ◽  
Author(s):  
Joni Frederick ◽  
Fritha Hennessy ◽  
Uli Horn ◽  
Pilar de la Torre Cortés ◽  
Marcel van den Broek ◽  
...  
Keyword(s):  
Nitrile Hydratase ◽  
Model Organism ◽  
Protein Sequences ◽  
Gene Clusters ◽  
Gram Positive Bacteria ◽  
Wide Range ◽  
Peptide Synthetase ◽  
Potential Applications ◽  
Rhodococcus Jostii Rha1 ◽  
Nitrile Hydrolysis

Abstract Background Rhodococci are industrially important soil-dwelling Gram-positive bacteria that are well known for both nitrile hydrolysis and oxidative metabolism of aromatics. Rhodococcus rhodochrous ATCC BAA-870 is capable of metabolising a wide range of aliphatic and aromatic nitriles and amides. The genome of the organism was sequenced and analysed in order to better understand this whole cell biocatalyst. Results The genome of R. rhodochrous ATCC BAA-870 is the first Rhodococcus genome fully sequenced using Nanopore sequencing. The circular genome contains 5.9 megabase pairs (Mbp) and includes a 0.53 Mbp linear plasmid, that together encode 7548 predicted protein sequences according to BASys annotation, and 5535 predicted protein sequences according to RAST annotation. The genome contains numerous oxidoreductases, 15 identified antibiotic and secondary metabolite gene clusters, several terpene and nonribosomal peptide synthetase clusters, as well as 6 putative clusters of unknown type. The 0.53 Mbp plasmid encodes 677 predicted genes and contains the nitrile converting gene cluster, including a nitrilase, a low molecular weight nitrile hydratase, and an enantioselective amidase. Although there are fewer biotechnologically relevant enzymes compared to those found in rhodococci with larger genomes, such as the well-known Rhodococcus jostii RHA1, the abundance of transporters in combination with the myriad of enzymes found in strain BAA-870 might make it more suitable for use in industrially relevant processes than other rhodococci. Conclusions The sequence and comprehensive description of the R. rhodochrous ATCC BAA-870 genome will facilitate the additional exploitation of rhodococci for biotechnological applications, as well as enable further characterisation of this model organism. The genome encodes a wide range of enzymes, many with unknown substrate specificities supporting potential applications in biotechnology, including nitrilases, nitrile hydratase, monooxygenases, cytochrome P450s, reductases, proteases, lipases, and transaminases.

scholarly journals Development of a tunable 3D wrinkled platfom for designing cellular three-dimensional communication networks

2021 ◽  
Author(s):  
Bethany R. Hughes
Keyword(s):  
Communication Networks ◽  
Three Dimensional ◽  
Cell Communication ◽  
Cellular Growth ◽  
Clinical Aspects ◽  
Wide Range ◽  
Cellular Communication Network ◽  
Potential Applications ◽  
A Cell ◽  
Washing Method

The study of cell-cell communication is hindered by the absence of a platform which is capable of specifically directing cellular growth while allowing examination of the communication between cells. In this thesis, a tuneable micro-to-nano scale wrinkled nonplanar platform was developed and optimized through the use of photolithography and a microfluidic washing method. The platform demonstrated the ability to create micro and nanowrinkled structures in a wide range of flow conditions. The developed platform was then used as a cell culture platform to investigate the spacing dependence of bovine fibroblasts. The identification of a critical bridging distance for bovine fibroblasts provided a means to optimized the platform for culturing a cellular communication network between bovine fibroblasts. The cellular network which resulted demonstrated, via FRAP (Fluorescence-recovery-after-photobleaching), the capacity for communication between cells. Creating multilevel length scaled structures on a tunable platform which directed cellular growth while maintaining communication presents potential applications in research, industry and clinical aspects.

Massive and Selective Ex Vivo Generation of Matured and Functional Human Red Blood Cells (RBC) from Hematopoietic Stem Cells of Diverse Origins: Towards the New Concept of “Cultured RBC”.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 379-379
Author(s):  
Luc Douay ◽  
Ladan Kobari ◽  
Helene Lapillonne ◽  
David Chalmers ◽  
Laurent Kiger ◽  
...  
Keyword(s):  
Large Scale ◽  
Ex Vivo ◽  
Erythroid Differentiation ◽  
Culture Conditions ◽  
Hematopoietic Stem ◽  
Human Red Blood Cells ◽  
Wide Range ◽  
Stromal Cell Line ◽  
A Cell ◽  
Free Media

Abstract We report a technological approach permitting, for the first time, the massive (up to 2x106-fold cell expansion) and selective (100%) ex vivo production of mature RBCs (cRBCs) starting from CD 34+ cells from peripheral blood (PB), bone marrow (BM) or cord blood (CB) into mature red cells in three steps: firstly, cell proliferation and erythroid differentiation were induced in serum free media supplemented with SCF, IL-3 and Epo for 8 days. Secondly, cells were co-cultured with additional Epo alone on either the murine MS-5 stromal cell line or human mesenchymal cells for 3 days. In the third step, all exogenous factors were withdrawn and cells were incubated on a simple stroma for 4 to 10 days. These cultured erythroid cells (reticulocytes and mature RBCs) displayed characteristics identical to those of native cells, in terms of MCV, MCH, MCHC, enzyme content (G6PD and PK) and deformability. The nature of the Hb produced depended on both the origin of the CD34+ cells and the culture conditions. cRBCs derived from PB or adult BM contained adult Hb (95±1%) whereas cRBCs derived from CB contained essentially HbF (64±13%). As for native RBCs, Hb was able to fix and release oxygen. CFSE-labelled-reticulocytes ex vivo generated from leukapheresis were injected into NOD-SCID mice. The transfused reticulocytes were found in the circulation to the same extent as native RBCs and fully matured into RBCs. This methodology is applicable for fundamental analysis of the mechanisms of terminal erythropoiesis and hemoglobin synthesis. Moreover, large scale cRBCs production could be possible with such a protocol. It can therefore be extrapolated to a wide range of clinical applications in the field of gene therapy, infectious diseases and particularly transfusion medicine with a pointed interest for the generation of a cell population homogeneous in age, thus achieving the new concept of cultured RBCs transfusion.

scholarly journals Supercapacitor-Assisted Techniques and Supercapacitor-Assisted Loss Management Concept: New Design Approaches to Change the Roadmap of Power Conversion Systems

Electronics ◽  
2021 ◽  
Vol 10 (14) ◽  
pp. 1697
Author(s):  
Nihal Kularatna ◽  
Kasun Subasinghage ◽  
Kosala Gunawardane ◽  
Dilini Jayananda ◽  
Thilanga Ariyarathna
Keyword(s):  
Power Conversion ◽  
Fold Increase ◽  
Voltage Regulation ◽  
Storage Unit ◽  
Energy Storage Unit ◽  
Low Dropout Regulators ◽  
Wide Range ◽  
A Cell ◽  
Main Component ◽  
Conversion Stage

All electrical and electronic devices require access to a suitable energy source. In a portable electronic product, such as a cell phone, an energy storage unit drives a complex array of power conversion stages to generate multiple DC voltage rails required. To optimize the overall end-to-end efficiency, these internal power conversions should waste minimal energy and deliver more to the electronic modules. Capacitors are one of the main component families used in electronics, to store and deliver electric charges. Supercapacitors, so called because they provide over a million-fold increase in capacitance relative to a traditional capacitor of the same volume, are enabling a paradigm shift in the design of power electronic converter circuits. Here we show that supercapacitors could function as a lossless voltage-dropping element in the power conversion stages, thereby significantly increasing the power conversion stage efficiency. This approach has numerous secondary benefits: it improves continuity of the supply, suppresses voltage surges, allows the voltage regulation to be electromagnetically silent, and simplifies the design of voltage regulators. The use of supercapacitors allows the development of a novel loss-circumvention theory with applicability to a wide range of supercapacitor-assisted (SCA) techniques. These include low-dropout regulators, transient surge absorbers, LED lighting for DC microgrids, and rapid energy transfer for water heating.

scholarly journals Improvement of an Escherichia coli whole-cell biocatalyst for geranyl glucoside production using directed evolution

2021 ◽  
Author(s):  
Julian Ruediger ◽  
Wilfried Schwab
Keyword(s):  
Directed Evolution ◽  
Production Systems ◽  
Low Cost ◽  
Continuous Production ◽  
Expression System ◽  
Screening Method ◽  
Fold Increase ◽  
Small Scale ◽  
Biotechnological Production ◽  
Whole Cell

The biotechnological production of glycosides is an economically competitive manufacturing alternative to classical chemical synthesis. Through continuous production improvement, glycosides can now be used in low-cost products by various industries. However, many production systems still suffer from low yields. Directed evolution, coupled with a suitable screening method, can tackle this challenge. We generated glycosyltransferase mutants through error-prone-PCR and screened the library using a small-scale whole-cell biotransformation system. The screening of only 176 colonies yielded three putative candidates. Detailed investigations revealed that the reason for the increase in product titer was mainly due to different expression effects of the mutagenized genes rather than improved enzyme kinetics. In total, a 60-fold increase in product formation was achieved. Therefore, in addition to the quality of the mutant library, an efficient and stable expression system is crucial to achieve high concentrations of active enzyme and product, as formation of inclusion bodies and other inactive forms of the biocatalyst reduces productivity.

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