Showing papers in "Applied Microbiology and Biotechnology in 2005"
TL;DR: There has been much industrial interest in xylan and its hydrolytic enzymatic complex, as a supplement in animal feed, for the manufacture of bread, food and drinks, textiles, bleaching of cellulose pulp, ethanol and xylitol production.
Abstract: Xylan is the principal type of hemicellulose. It is a linear polymer of beta-D-xylopyranosyl units linked by (1-4) glycosidic bonds. In nature, the polysaccharide backbone may be added to 4-O-methyl-alpha-D-glucuronopyranosyl units, acetyl groups, alpha-L-arabinofuranosyl, etc., in variable proportions. An enzymatic complex is responsible for the hydrolysis of xylan, but the main enzymes involved are endo-1,4-beta-xylanase and beta-xylosidase. These enzymes are produced by fungi, bacteria, yeast, marine algae, protozoans, snails, crustaceans, insect, seeds, etc., but the principal commercial source is filamentous fungi. Recently, there has been much industrial interest in xylan and its hydrolytic enzymatic complex, as a supplement in animal feed, for the manufacture of bread, food and drinks, textiles, bleaching of cellulose pulp, ethanol and xylitol production. This review describes some properties of xylan and its metabolism, as well as the biochemical properties of xylanases and their commercial applications.
1,214 citations
TL;DR: The algal biodiversity, the chemical structures and biosynthesis of hydrocarbons and ether lipids and the biotechnological studies related to hydrocarbon production are reviewed.
Abstract: This paper presents a review on Botryococcus braunii, a cosmopolitan green colonial microalga characterised by a considerable production of lipids, notably hydrocarbons. Strains like wild populations of this alga differ in the type of hydrocarbons they synthesise and accumulate: (1) n-alkadienes and trienes, (2) triterpenoid botryococcenes and methylated squalenes, or (3) a tetraterpenoid, lycopadiene. In addition to hydrocarbons and some classic lipids, these algae produce numerous series of characteristic ether lipids closely related to hydrocarbons. This review covers the algal biodiversity, the chemical structures and biosynthesis of hydrocarbons and ether lipids and the biotechnological studies related to hydrocarbon production.
734 citations
TL;DR: For almost 50 years now, biotechnological production processes have been used for industrial production of amino acids, and the use of enzymes and whole cell biocatalysts has proven particularly valuable in production of both proteinogenic and nonproteinogenic l-amino acids.
Abstract: For almost 50 years now, biotechnological production processes have been used for industrial production of amino acids. Market development has been particularly dynamic for the flavor-enhancer glutamate and the animal feed amino acids L: -lysine, L: -threonine, and L: -tryptophan, which are produced by fermentation processes using high-performance strains of Corynebacterium glutamicum and Escherichia coli from sugar sources such as molasses, sucrose, or glucose. But the market for amino acids in synthesis is also becoming increasingly important, with annual growth rates of 5-7%. The use of enzymes and whole cell biocatalysts has proven particularly valuable in production of both proteinogenic and nonproteinogenic L: -amino acids, D: -amino acids, and enantiomerically pure amino acid derivatives, which are of great interest as building blocks for active ingredients that are applied as pharmaceuticals, cosmetics, and agricultural products. Nutrition and health will continue to be the driving forces for exploiting the potential of microorganisms, and possibly also of suitable plants, to arrive at even more efficient processes for amino acid production.
723 citations
TL;DR: While fuel ethanol production is an energy-efficient process today, additional research is occurring to improve its long-term economic viability and three of the most significant areas of research are in the production of hybrids with a higher starch content or a higher extractable starch content, in the conversion of the corn kernel fiber fraction to ethanol, and in the identification and development of new and higher-value co-products.
Abstract: Ethanol has been utilized as a fuel source in the United States since the turn of the century. However, it has repeatedly faced significant commercial viability obstacles relative to petroleum. Renewed interest exists in ethanol as a fuel source today owing to its positive impact on rural America, the environment and United States energy security. Today, most fuel ethanol is produced by either the dry grind or the wet mill process. Current technologies allow for 2.5 gallons (wet mill process) to 2.8 gallons (dry grind process) of ethanol (1 gallon = 3.785 l) per bushel of corn. Valuable co-products, distillers dried grains with solubles (dry grind) and corn gluten meal and feed (wet mill), are also generated in the production of ethanol. While current supplies are generated from both processes, the majority of the growth in the industry is from dry grind plant construction in rural communities across the corn belt. While fuel ethanol production is an energy-efficient process today, additional research is occurring to improve its long-term economic viability. Three of the most significant areas of research are in the production of hybrids with a higher starch content or a higher extractable starch content, in the conversion of the corn kernel fiber fraction to ethanol, and in the identification and development of new and higher-value co-products.
664 citations
TL;DR: An extensive overview is provided of the different applications in lactococci and other Gram-positive bacteria: over-expression of homologous and heterologous genes for functional studies and to obtain large quantities of specific gene products, metabolic engineering, and large-scale applications.
Abstract: Lactococcus lactis is a Gram-positive lactic acid bacterium that, in addition to its traditional use in food fermentations, is increasingly used in modern biotechnological applications. In the last 25 years great progress has been made in the development of genetic engineering tools and the molecular characterization of this species. A new versatile and tightly controlled gene expression system, based on the auto-regulation mechanism of the bacteriocin nisin, was developed 10 years ago-the NIsin Controlled gene Expression system, called NICE. This system has become one of the most successful and widely used tools for regulated gene expression in Gram-positive bacteria. The review describes, after a brief introduction of the host bacterium L. lactis, the fundaments, components and function of the NICE system. Furthermore, an extensive overview is provided of the different applications in lactococci and other Gram-positive bacteria: (1) over-expression of homologous and heterologous genes for functional studies and to obtain large quantities of specific gene products, (2) metabolic engineering, (3) expression of prokaryotic and eukaryotic membrane proteins, (4) protein secretion and anchoring in the cell envelope, (5) expression of genes with toxic products and analysis of essential genes and (6) large-scale applications. Finally, an overview is given of growth and induction conditions for lab-scale and industrial-scale applications.
542 citations
TL;DR: The ability to produce the ever-increasing number of biopharmaceuticals by animal cell culture is dependent on sufficient bioreactor capacity in the industry, and a recent shortfall in available worldwide culture capacity has encouraged commercial activity in contract manufacturing operations.
Abstract: There has been a rapid increase in the number and demand for approved biopharmaceuticals produced from animal cell culture processes over the last few years. In part, this has been due to the efficacy of several humanized monoclonal antibodies that are required at large doses for therapeutic use. There have also been several identifiable advances in animal cell technology that has enabled efficient biomanufacture of these products. Gene vector systems allow high specific protein expression and some minimize the undesirable process of gene silencing that may occur in prolonged culture. Characterization of cellular metabolism and physiology has enabled the design of fed-batch and perfusion bioreactor processes that has allowed a significant improvement in product yield, some of which are now approaching 5 g/L. Many of these processes are now being designed in serum-free and animal-component-free media to ensure that products are not contaminated with the adventitious agents found in bovine serum. There are several areas that can be identified that could lead to further improvement in cell culture systems. This includes the down-regulation of apoptosis to enable prolonged cell survival under potentially adverse conditions. The characterization of the critical parameters of glycosylation should enable process control to reduce the heterogeneity of glycoforms so that production processes are consistent. Further improvement may also be made by the identification of glycoforms with enhanced biological activity to enhance clinical efficacy. The ability to produce the ever-increasing number of biopharmaceuticals by animal cell culture is dependent on sufficient bioreactor capacity in the industry. A recent shortfall in available worldwide culture capacity has encouraged commercial activity in contract manufacturing operations. However, some analysts indicate that this still may not be enough and that future manufacturing demand may exceed production capacity as the number of approved biotherapeutics increases.
521 citations
TL;DR: It is suggested that active inoculating techniques could increase the effectiveness of enrichment, and that start up is most successful when the biofilm is harvested from the anode of an existing MFC and applied to the new anode.
Abstract: A microbial fuel cell (MFC) is a relatively new type of fixed film bioreactor for wastewater treatment, and the most effective methods for inoculation are not well understood. Various techniques to enrich electrochemically active bacteria on an electrode were therefore studied using anaerobic sewage sludge in a two-chambered MFC. With a porous carbon paper anode electrode, 8 mW/m2 of power was generated within 50 h with a Coulombic efficiency (CE) of 40%. When an iron oxide-coated electrode was used, the power and the CE reached 30 mW/m2 and 80%, respectively. A methanogen inhibitor (2-bromoethanesulfonate) increased the CE to 70%. Bacteria in sludge were enriched by serial transfer using a ferric iron medium, but when this enrichment was used in a MFC the power was lower (2 mW/m2) than that obtained with the original inoculum. By applying biofilm scraped from the anode of a working MFC to a new anode electrode, the maximum power was increased to 40 mW/m2. When a second anode was introduced into an operating MFC the acclimation time was not reduced and the total power did not increase. These results suggest that these active inoculating techniques could increase the effectiveness of enrichment, and that start up is most successful when the biofilm is harvested from the anode of an existing MFC and applied to the new anode.
501 citations
TL;DR: The rhamnolipids biosynthetic pathway has metabolic links with numerous bacterial products such as alginate, lipopolysaccharide, polyhydroxyalkanoates, and 4-hydroxy-2-alkylquinolines (HAQs).
Abstract: Pseudomonas aeruginosa produces glycolipidic surface-active molecules (rhamnolipids) which have potential biotechnological applications. Rhamnolipids are produced by P. aeruginosa in a concerted manner with different virulence-associated traits. Here, we review the rhamnolipids biosynthetic pathway, showing that it has metabolic links with numerous bacterial products such as alginate, lipopolysaccharide, polyhydroxyalkanoates, and 4-hydroxy-2-alkylquinolines (HAQs). We also discuss the factors controlling the production of rhamnolipids and the proposed roles this biosurfactant plays in P. aeruginosa lifestyle.
421 citations
TL;DR: High-molecular-weight polysaccharide–protein complexes from mushrooms that appear to enhance innate and cell-mediated immune responses, exhibit antitumor activities in animals and humans, and demonstrate the anticancer properties of selenium compounds accumulated in mushrooms are discussed.
Abstract: Empirical approaches to discover anticancer drugs and cancer treatments have made limited progress in the past several decades in finding a cure for cancer. The expanded knowledge of the molecular basis of tumorigenesis and metastasis, together with the inherently vast structural diversity of natural compounds found in mushrooms, provided unique opportunities for discovering new drugs that rationally target the abnormal molecular and biochemical signals leading to cancer. This review focuses on mushroom low-molecular-weight secondary metabolites targeting processes such as apoptosis, angiogenesis, metastasis, cell cycle regulation, and signal transduction cascades. Also discussed in this review are high-molecular-weight polysaccharides or polysaccharide-protein complexes from mushrooms that appear to enhance innate and cell-mediated immune responses, exhibit antitumor activities in animals and humans, and demonstrate the anticancer properties of selenium compounds accumulated in mushrooms.
393 citations
TL;DR: This review includes information on the structure, properties and molecular genetics of the bacterial (1→3)-β-glucans, together with an overview of the physiology and biotechnology of curdlan production and applications of this biopolymer and its derivatives.
Abstract: Three structural classes of (1-->3)-beta-D-glucans are encountered in some important soil-dwelling, plant-associated or human pathogenic bacteria. Linear (1-->3)-beta-glucans and side-chain-branched (1-->3,1-->2)-beta-glucans are major constituents of capsular materials, with roles in bacterial aggregation, virulence and carbohydrate storage. Cyclic (1-->3,1-->6)-beta-glucans are predominantly periplasmic, serving in osmotic adaptation. Curdlan, the linear (1-->3)-beta-glucan from Agrobacterium, has unique rheological and thermal gelling properties, with applications in the food industry and other sectors. This review includes information on the structure, properties and molecular genetics of the bacterial (1-->3)-beta-glucans, together with an overview of the physiology and biotechnology of curdlan production and applications of this biopolymer and its derivatives.
387 citations
TL;DR: In mineral salts medium under oxygen deprivation, Corynebacterium glutamicum exhibits high productivity of l-lactic acid accompanied with succinic and acetic acids, and was genetically modified to produce d-lactate dehydrogenase-encoding genes from Escherichia coli and Lactobacillus delbrueckii.
Abstract: In mineral salts medium under oxygen deprivation, Corynebacterium glutamicum exhibits high productivity of l-lactic acid accompanied with succinic and acetic acids. In taking advantage of this elevated productivity, C. glutamicum was genetically modified to produce d-lactic acid. The modification involved expression of fermentative d-lactate dehydrogenase (d-LDH)-encoding genes from Escherichia coli and Lactobacillus delbrueckii in l-lactate dehydrogenase (l-LDH)-encoding ldhA-null C. glutamicum mutants to yield strains C. glutamicum ΔldhA/pCRB201 and C. glutamicum ΔldhA/pCRB204, respectively. The productivity of C. glutamicum ΔldhA/pCRB204 was fivefold higher than that of C. glutamicum ΔldhA/pCRB201. By using C. glutamicum ΔldhA/pCRB204 cells packed to a high density in mineral salts medium, up to 1,336 mM (120 g l−1) of d-lactic acid of greater than 99.9% optical purity was produced within 30 h.
TL;DR: A broad set of bacterial strategies to degrade chlorobenzoates has recently been elucidated, including new pathways for the degradation of chlorocatechols as central intermediates of various chloroaromatic catabolic pathways.
Abstract: The microbial degradation of polychlorinated biphenyls (PCBs) has been extensively studied in recent years The genetic organization of biphenyl catabolic genes has been elucidated in various groups of microorganisms, their structures have been analyzed with respect to their evolutionary relationships, and new information on mobile elements has become available Key enzymes, specifically biphenyl 2,3-dioxygenases, have been intensively characterized, structure/sequence relationships have been determined and enzymes optimized for PCB transformation However, due to the complex metabolic network responsible for PCB degradation, optimizing degradation by single bacterial species is necessarily limited As PCBs are usually not mineralized by biphenyl-degrading organisms, and cometabolism can result in the formation of toxic metabolites, the degradation of chlorobenzoates has received special attention A broad set of bacterial strategies to degrade chlorobenzoates has recently been elucidated, including new pathways for the degradation of chlorocatechols as central intermediates of various chloroaromatic catabolic pathways To optimize PCB degradation in the environment beyond these metabolic limitations, enhancing degradation in the rhizosphere has been suggested, in addition to the application of surfactants to overcome bioavailability barriers However, further research is necessary to understand the complex interactions between soil/sediment, pollutant, surfactant and microorganisms in different environments
TL;DR: Metabolic engineering is providing new opportunities and HA produced in a heterologous host is about to enter the market, but greater understanding of the mechanisms underlying chain termination is required.
Abstract: Hyaluronic acid (HA) is a commercially valuable medical biopolymer increasingly produced through microbial fermentation. Viscosity limits product yield and the focus of research and development has been on improving the key quality parameters, purity and molecular weight. Traditional strain and process optimisation has yielded significant improvements, but appears to have reached a limit. Metabolic engineering is providing new opportunities and HA produced in a heterologous host is about to enter the market. In order to realise the full potential of metabolic engineering, however, greater understanding of the mechanisms underlying chain termination is required.
TL;DR: This is the first significant effort to achieve an electronic display using bacterial cellulose and has the potential to be extended to various applications, such as e-book tablets, e-newspapers, dynamic wall papers, rewritable maps and learning tools.
Abstract: Cellulose (in the form of printed paper) has always been the prime medium for displaying information in our society and is far better than the various existing display technologies. This is because of its high reflectivity, contrast, low cost and flexibility. There is a major initiative to push for a dynamic display technology that emulates paper (popularly known as “electronic paper”). We have successfully demonstrated the proof of the concept of developing a dynamic display on cellulose. To the best of our knowledge, this is the first significant effort to achieve an electronic display using bacterial cellulose. First, bacterial cellulose is synthesized in a culture of Acetobacter xylinum in standard glucose-rich medium. The bacterial cellulose membrane thus formed (not pulp) is dimensionally stable, has a paper-like appearance and has a unique microfibrillar nanostructure. The technique then involves first making the cellulose an electrically conducting (or semi-conducting) sheet by depositing ions around the microfibrils to provide conducting pathways and then immobilizing electrochromic dyes within the microstructure. The whole system is then cased between transparent electrodes, and upon application of switching potentials (2–5 V) a reversible color change can be demonstrated down to a standard pixel-sized area (ca. 100 μm2). Using a standard back-plane or in-plane drive circuit, a high-resolution dynamic display device using cellulose as substrate can be constructed. The major advantages of such a device are its high paper-like reflectivity, flexibility, contrast and biodegradability. The device has the potential to be extended to various applications, such as e-book tablets, e-newspapers, dynamic wall papers, rewritable maps and learning tools.
TL;DR: Antimicrobial berry compounds may have important applications in the future as natural antimicrobial agents for food industry as well as for medicine.
Abstract: Berry fruits are rich sources of bioactive compounds, such as phenolics and organic acids, which have antimicrobial activities against human pathogens. Among different berries and berry phenolics, cranberry, cloudberry, raspberry, strawberry and bilberry especially possess clear antimicrobial effects against, e.g. Salmonella and Staphylococcus. Complex phenolic polymers, like ellagitannins, are strong antibacterial agents present in cloudberry and raspberry. Several mechanisms of action in the growth inhibition of bacteria are involved, such as destabilisation of cytoplasmic membrane, permeabilisation of plasma membrane, inhibition of extracellular microbial enzymes, direct actions on microbial metabolism and deprivation of the substrates required for microbial growth. Antimicrobial activity of berries may also be related to antiadherence of bacteria to epithelial cells, which is a prerequisite for colonisation and infection of many pathogens. Antimicrobial berry compounds may have important applications in the future as natural antimicrobial agents for food industry as well as for medicine. Some of the novel approaches are discussed.
TL;DR: This review summarizes recent developments and demonstrates that aptamers are valuable tools for diagnostics, purification processes, target validation, drug discovery, and even therapeutic approaches.
Abstract: Since its discovery in the early 1990s, aptamer technology has progressed tremendously. Automated selection procedures now allow rapid identification of DNA and RNA sequences that can target a broad range of extra- and intracellular proteins with nanomolar affinities and high specificities. The unique binding properties of nucleic acids, which are amenable to various modifications, make aptamers perfectly suitable for different areas of biotechnology. Moreover, the approval of an aptamer for vascular endothelial growth factor by the US Food and Drug Administration highlights the potential of aptamers for therapeutic applications. This review summarizes recent developments and demonstrates that aptamers are valuable tools for diagnostics, purification processes, target validation, drug discovery, and even therapeutic approaches.
TL;DR: Evidence is provided for the role of secreted lipopeptides, and more particularly of fengycins, in the protective effect afforded by the strain against damping-off of bean seedlings caused by Pythium ultimum and against gray mold of apple in post-harvest disease.
Abstract: In this work, the potential of Bacillus subtilis strain M4 at protecting plants against fungal diseases was demonstrated in different pathosystems. We provide evidence for the role of secreted lipopeptides, and more particularly of fengycins, in the protective effect afforded by the strain against damping-off of bean seedlings caused by Pythium ultimum and against gray mold of apple in post-harvest disease. This role was demonstrated by the strong biocontrol activity of lipopeptide-enriched extracts and through the detection of inhibitory quantities of fengycins in infected tissues. Beside such a direct antagonism of the pathogen, we show that root pre-inoculation with M4 enabled the host plant to react more efficiently to subsequent pathogen infection on leaves. Fengycins could also be involved in this systemic resistance-eliciting effect of strain M4, as these molecules may induce the synthesis of plant phenolics involved in or derived from the defense-related phenylpropanoid metabolism. Much remains to be discovered about the mechanisms by which Bacillus spp suppress disease. Through this study on strain M4, we reinforce the interest in B. subtilis as a pathogen antagonist and plant defense-inducing agent. The secretion of cyclic fengycin-type lipopeptides may be tightly related to the expression of these two biocontrol traits.
TL;DR: The potential applications of esterase with reference to agriculture, food, and pharmaceutical industries, are discussed in this review.
Abstract: Esterase plays a major role in the degradation of natural materials and industrial pollutants, viz, cereal wastes, plastics, and other toxic chemicals It is useful in the synthesis of optically pure compounds, perfumes, and antioxidants The potential applications of esterase with reference to agriculture, food, and pharmaceutical industries, are discussed in this review Promising applications in this avenue can be supported by appropriate production strategies
TL;DR: In addition to its major application in animal nutrition, phytase is also used for processing of human food and research in this field focuses on better mineral absorption and technical improvement of food processing.
Abstract: Phytases decompose phytate, which is the primary storage form of phosphate in plants. More than 10 years ago, the first commercial phytase product became available on the market. It offered to help farmers reduce phosphorus excretion of monogastric animals by replacing inorganic phosphates by microbial phytase in the animal diet. Phytase application can reduce phosphorus excretion by up to 50%, a feat that would contribute significantly toward environmental protection. Furthermore, phytase supplementation leads to improved availability of minerals and trace elements. In addition to its major application in animal nutrition, phytase is also used for processing of human food. Research in this field focuses on better mineral absorption and technical improvement of food processing. All commercial phytase preparations contain microbial enzymes produced by fermentation. A wide variety of phytases were discovered and characterized in the last 10 years. Initial steps to produce phytase in transgenic plants were also undertaken. A crucial role for its commercial success relates to the formulation of the enzyme solution delivered from fermentation. For liquid enzyme products, a long shelf life is achieved by the addition of stabilizing agents. More comfortable for many customers is the use of dry enzyme preparations. Different formulation technologies are used to produce enzyme powders that retain enzyme activity, are stable in application, resistant against high temperatures, dust-free, and easy to handle.
TL;DR: The importance of monoterpenes on varietal flavour of must and other fruit juices has been reviewed and analytical techniques developed to study these compounds, in both free or glycosidically forms, are discussed.
Abstract: The importance of monoterpenes on varietal flavour of must and other fruit juices has been reviewed. These compounds were mainly found linked to sugar moieties in grape juice and wines, showing no olfactory characteristics. In this way, analytical techniques developed to study these compounds, in both free or glycosidically forms, are discussed. Mechanisms to liberate terpenes were studied, making a comparative study between acidic and enzymic hydrolysis of terpene glycosides; as enzymic hydrolysis seems to be the most natural way to liberate terpenes, the ability to use glycosidases from grapes, yeasts, bacterial or exogenous, i.e. fungal commercial preparations, were reviewed. Re-arrangements of terpenes after acidic hydrolysis of glycoconjugated are discussed as well as potential adverse effects of enzyme preparations.
TL;DR: An overview on the problems typically coming along with fermentation process optimization and scale up is given, and currently applied scale-up strategies while considering future technologies are presented, with emphasis on Escherichia coli as one of the most commonly fermented organisms.
Abstract: To increase product yields and to ensure consistent product quality, key issues of industrial fermentations, process optimization and scale up are aimed at maintaining optimum and homogenous reaction conditions minimizing microbial stress exposure and enhancing metabolic accuracy. For each individual product, process and facility, suitable strategies have to be elaborated by a comprehensive and detailed process characterization, identification of the most relevant process parameters influencing product yield and quality and their establishment as scale-up parameters to be kept constant as far as possible. Physical variables, which can only be restrictedly kept constant as single parameters, may be combined with other pertinent parameters to appropriate mathematical groups or dimensionless terms. Process characterization is preferably based on real-time or near real-time data collected by in situ and on-line measurements and may be facilitated by supportive approaches and tools like neural network based chemometric data analysis and modelling, clarification of the mixing and stream conditions through computational fluid dynamics and scale-down simulations. However, as fermentation facilities usually are not strictly designed according to scale-up criteria and the process conditions in the culture vessels thus may differ significantly and since any strategy and model can only insufficiently consider and reflect the highly complex interdependence and mutual interaction of fermentation parameters, successful scale up in most cases is not the result of a conclusive and straight-lined experimental strategy, but rather will be the outcome of a separate process development and optimization on each scale. This article gives an overview on the problems typically coming along with fermentation process optimization and scale up, and presents currently applied scale-up strategies while considering future technologies, with emphasis on Escherichia coli as one of the most commonly fermented organisms.
TL;DR: In recent years, the number of recombinant proteins used for therapeutic applications has increased dramatically, and production of these proteins has a remarkable demand in the market, with major challenges is obtaining the high yield of protein at low cost.
Abstract: In recent years, the number of recombinant proteins used for therapeutic applications has increased dramatically. Production of these proteins has a remarkable demand in the market. Escherichia coli offers a means for the rapid and economical production of recombinant proteins. These advantages, coupled with a wealth of biochemical and genetic knowledge, have enabled the production of such economically therapeutic proteins such as insulin and bovine growth hormone. These demands have driven the development of a variety of strategies for achieving high-level expression of protein, particularly involving several aspects such as expression vectors design, gene dosage, promoter strength (transcriptional regulation), mRNA stability, translation initiation and termination (translational regulation), host design considerations, codon usage, and fermentation factors available for manipulating the expression conditions, which are the major challenges is obtaining the high yield of protein at low cost.
TL;DR: This cell-recycling continuous reaction demonstrated that rates of organic acid production by C. glutamicum could be maintained for at least 360 h, and there was a direct correlation between cell concentration and organic acids production rates even at elevated cell densities.
Abstract: Under oxygen deprivation, aerobic Corynebacterium glutamicum produce organic acids from glucose at high yields in mineral medium even though their proliferation is arrested. To develop a new, high-productivity bioprocess based on these unique features, characteristics of organic acid production by C. glutamicum under oxygen deprivation were investigated. The main organic acids produced from glucose under these conditions were lactic acid and succinic acid. Addition of bicarbonate, which is a co-substrate for anaplerotic enzymes, increased the glucose consumption rate, leading to increased organic acid production rates. With increasing concentration of bicarbonate, the yield of succinic acid increased, whereas that of lactic acid decreased. There was a direct correlation between cell concentration and organic acid production rates even at elevated cell densities, and productivities of lactic acid and succinic acid were 42.9 g l−1 h−1 and 11.7 g l−1 h−1, respectively, at a cell concentration of 60 g dry cell l−1. This cell-recycling continuous reaction demonstrated that rates of organic acid production by C. glutamicum could be maintained for at least 360 h.
TL;DR: Aerobic granular sludge can successfully be cultivated in a sequencing batch reactor (SBR) treating dairy wastewater, and the maximum applicable loading rate is nevertheless limited, as the stability of aerobic granules very much depends on the presence of distinct feast and famine conditions.
Abstract: Aerobic granular sludge can successfully be cultivated in a sequencing batch reactor (SBR) treating dairy wastewater. Attention has to be paid to the fact that suspended solids are always present in the effluent of aerobic granular sludge reactors, making a post-treatment step necessary. Sufficient post-treatment can be achieved through a sedimentation process with a hydraulic retention time of 15–30 min. After complete granulation and the separation of biomass from the effluent, removal efficiencies of 90% CODtotal, 80% Ntotal and 67% Ptotal can be achieved at a volumetric exchange ratio of 50% and a cycle duration of 8 h. Effluent values stabilize at around 125 mg l−1 CODdissolved. The maximum applicable loading rate is nevertheless limited, as the stability of aerobic granules very much depends on the presence of distinct feast and famine conditions and the degradation of real wastewaters shows slower kinetics compared with synthetic wastewaters. As loading rate and volumetric exchange ratio are coupled in an SBR system, the potential of granular sludge for improving process efficiency is also limited.
TL;DR: In this article, the fibrinolytic enzymes were successively discovered from different microorganisms, the most important among which is the genus Bacillus from traditional fermented foods, and the physiochemical properties of these enzymes have been characterized, and their effectiveness in thrombolysis in vivo has been further identified.
Abstract: Accumulation of fibrin in the blood vessels usually results in thrombosis, leading to myocardial infarction and other cardiovascular diseases. For thrombolytic therapy, microbial fibrinolytic enzymes have now attracted much more attention than typical thrombolytic agents because of the expensive prices and the undesirable side effects of the latter. The fibrinolytic enzymes were successively discovered from different microorganisms, the most important among which is the genus Bacillus from traditional fermented foods. The physiochemical properties of these enzymes have been characterized, and their effectiveness in thrombolysis in vivo has been further identified. Therefore, microbial fibrinolytic enzymes, especially those from food-grade microorganisms, have the potential to be developed as functional food additives and drugs to prevent or cure thrombosis and other related diseases.
TL;DR: Early approaches to model this morphologic development are recapitulated in this review to explain current trends in this area of research and present recent advances in the investigation of fungal metabolism.
Abstract: Cultivation processes involving filamentous fungi have been optimised for decades to obtain high product yields. Several bulk chemicals like citric acid and penicillin are produced this way. A simple adaptation of cultivation parameters for new production processes is not possible though. Models explaining the correlation between process-dependent growth behaviour and productivity are therefore necessary to prevent long-lasting empiric test series. Yet, filamentous growth consists of a complex microscopic differentiation process from conidia to hyphae resulting in various macroscopically visible appearances. Early approaches to model this morphologic development are recapitulated in this review to explain current trends in this area of research. Tailoring morphology by adjusting process parameters is one side of the coin, but an ideal morphology has not even been found. This article reviews several reasons for this fact starting with nutrient supply in a fungal culture and presents recent advances in the investigation of fungal metabolism. It illustrates the challenge to unfold the relationship between morphology and productivity.
TL;DR: This study demonstrates that, using effective annotation tools, genomes can efficiently be exploited as a source of novel BVMOs, and identifies a newly identified biocatalyst that is relatively thermostable and able to perform sulfur oxidations.
Abstract: Baeyer–Villiger monooxygenases represent useful biocatalytic tools, as they can catalyze reactions which are difficult to achieve using chemical means. However, only a limited number of these atypical monooxygenases are available in recombinant form. Using a recently described protein sequence motif, a putative Baeyer–Villiger monooxygenase (BVMO) was identified in the genome of the thermophilic actinomycete Thermobifida fusca. Heterologous expression of the respective protein in Escherichia coli and subsequent enzyme characterization showed that it indeed represents a BVMO. The NADPH-dependent and FAD-containing monooxygenase is active with a wide range of aromatic ketones, while aliphatic substrates are also converted. The best substrate discovered so far is phenylacetone (kcat = 1.9 s−1, KM = 59 μM). The enzyme exhibits moderate enantioselectivity with α-methylphenylacetone (enantiomeric ratio of 7). In addition to Baeyer–Villiger reactions, the enzyme is able to perform sulfur oxidations. Different from all known BVMOs, this newly identified biocatalyst is relatively thermostable, displaying an activity half-life of 1 day at 52°C. This study demonstrates that, using effective annotation tools, genomes can efficiently be exploited as a source of novel BVMOs.
TL;DR: In recent years, the number of recombinant proteins used for therapeutic applications has increased dramatically, and production of these proteins has a remarkable demand in the market, with major challenges is obtaining the high yield of protein at low cost.
Abstract: In recent years, the number of recombinant proteins used for therapeutic applications has increased dramatically. Production of these proteins has a remarkable demand in the market. Escherichia coli offers a means for the rapid and economical production of recombinant proteins. These advantages, coupled with a wealth of biochemical and genetic knowledge, have enabled the production of such economically therapeutic proteins such as insulin and bovine growth hormone. These demands have driven the development of a variety of strategies for achieving high-level expression of protein, particularly involving several aspects such as expression vectors design, gene dosage, promoter strength (transcriptional regulation), mRNA stability, translation initiation and termination (translational regulation), host design considerations, codon usage, and fermentation factors available for manipulating the expression conditions, which are the major challenges is obtaining the high yield of protein at low cost.
TL;DR: Determination of the affinities for different chemicals indicated that the most preferred substrate for the enzyme would consist of butyric acid and n-hexanol, and the environment of the catalytic pocket, which is divided into carboxylic acid and alcohol recognition sites, was investigated.
Abstract: We used biodegradable plastics as fermentation substrates for the filamentous fungus Aspergillus oryzae. This fungus could grow under culture conditions that contained emulsified poly-(butylene succinate) (PBS) and emulsified poly-(butylene succinate-co-adipate) (PBSA) as the sole carbon source, and could digest PBS and PBSA, as indicated by clearing of the culture supernatant. We purified the PBS-degrading enzyme from the culture supernatant, and its molecular mass was determined as 21.6 kDa. The enzyme was identified as cutinase based on internal amino acid sequences. Specific activities against PBS, PBSA and poly-(lactic acid) (PLA) were determined as 0.42 U/mg, 11 U/mg and 0.067 U/mg, respectively. To obtain a better understanding of how the enzyme recognizes and hydrolyzes PBS/PBSA, we investigated the environment of the catalytic pocket, which is divided into carboxylic acid and alcohol recognition sites. The affinities for different substrates depended on the carbon chain length of the carboxylic acid in the substrate. Competitive inhibition modes were exhibited by carboxylic acids and alcohols that consisted of C4-C6 and C3-C8 chain lengths, respectively. Determination of the affinities for different chemicals indicated that the most preferred substrate for the enzyme would consist of butyric acid and n-hexanol.
TL;DR: UV–visible spectrum analysis, an easy and rapid technique, could be used to monitor cell lysis during EPS extraction from R. acidophila and suggests that few cells were destroyed and lysis did not occur.
Abstract: Among the four methods for extracting extracellular polymeric substances (EPS) from Rhodopseudomonas acidophila (EDTA, NaOH, H(2)SO(4), heating/centrifugation), EDTA extraction was found to be the most effective. The contents of the major components of EPS from R. acidophila, i.e., carbohydrate, protein and nucleic acid, were 6.5, 58.4 and 5.4 mg g(-1) dry cells, respectively. The optimum extraction time was 1-3 h and the EDTA dosage was approximately 2.8 g g(-1) dry cells. Under these conditions, no cell lysis was observed. The EPS content and the percentage of the three main components were greatly dependent on the extraction method. The intensity of absorption peaks for photosynthetic pigments in the UV-visible spectrum of bacteria remained unchanged prior to and after EDTA extraction; and no pigment peaks appeared in the EPS spectrum. This suggests that few cells were destroyed and lysis did not occur. UV-visible spectrum analysis, an easy and rapid technique, could be used to monitor cell lysis during EPS extraction from R. acidophila.