Showing papers in "Applied Microbiology and Biotechnology in 2006"
TL;DR: The prospects included are fermentation technology converting xylose to ethanol, cellulase enzyme utilized in the hydrolysis of lignocellulosic materials, immobilization of the microorganism in large systems, simultaneous saccharification and fermentation, and sugar conversion into ethanol.
Abstract: In recent years, growing attention has been devoted to the conversion of biomass into fuel ethanol, considered the cleanest liquid fuel alternative to fossil fuels. Significant advances have been made towards the technology of ethanol fermentation. This review provides practical examples and gives a broad overview of the current status of ethanol fermentation including biomass resources, microorganisms, and technology. Also, the promising prospects of ethanol fermentation are especially introduced. The prospects included are fermentation technology converting xylose to ethanol, cellulase enzyme utilized in the hydrolysis of lignocellulosic materials, immobilization of the microorganism in large systems, simultaneous saccharification and fermentation, and sugar conversion into ethanol.
1,610 citations
TL;DR: An overview of the most commonly used promoter systems for recombinant proteins, including Bacillus brevis, Bacillusmegaterium, Bacillussubtilis, Caulobacter crescentus, other strains, and, most importantly, Escherichia coli BL21 and E. coli K12 and their derivatives are presented.
Abstract: During the proteomics period, the growth in the use of recombinant proteins has increased greatly in the recent years. Bacterial systems remain most attractive due to low cost, high productivity, and rapid use. However, the rational choice of the adequate promoter system and host for a specific protein of interest remains difficult. This review gives an overview of the most commonly used systems: As hosts, Bacillus brevis, Bacillusmegaterium, Bacillussubtilis, Caulobacter crescentus, other strains, and, most importantly, Escherichia coli BL21 and E. coli K12 and their derivatives are presented. On the promoter side, the main features of the l-arabinose inducible araBAD promoter (PBAD), the lac promoter, the l-rhamnose inducible rhaPBAD promoter, the T7 RNA polymerase promoter, the trc and tac promoter, the lambda phage promoter pL, and the anhydrotetracycline-inducible tetA promoter/operator are summarized.
985 citations
TL;DR: This review describes a brief overview of the current research worldwide on the use of microorganisms in the biosynthesis of metal nanoparticles and their applications.
Abstract: Nanomaterials are at the leading edge of the rapidly developing field of nanotechnology. The development of reliable experimental protocols for the synthesis of nanomaterials over a range of chemical compositions, sizes, and high monodispersity is one of the challenging issues in current nanotechnology. In the context of the current drive to develop green technologies in material synthesis, this aspect of nanotechnology is of considerable importance. Biological systems, masters of ambient condition chemistry, synthesize inorganic materials that are hierarchically organized from the nano- to the macroscale. Recent studies on the use of microorganisms in the synthesis of nanoparticles are a relatively new and exciting area of research with considerable potential for development. This review describes a brief overview of the current research worldwide on the use of microorganisms in the biosynthesis of metal nanoparticles and their applications.
926 citations
TL;DR: This review provides the current status in the research of proteolytic systems of LAB with industrial relevance and indicates that the current knowledge on stress-related proteolysis in LAB is almost exclusively based on studies on L. lactis.
Abstract: Lactic acid bacteria (LAB) have a very long history of use in the manufacturing processes of fermented foods and a great deal of effort was made to investigate and manipulate the role of LAB in these processes. Today, the diverse group of LAB includes species that are among the best-studied microorganisms and proteolysis is one of the particular physiological traits of LAB of which detailed knowledge was obtained. The proteolytic system involved in casein utilization provides cells with essential amino acids during growth in milk and is also of industrial importance due to its contribution to the development of the organoleptic properties of fermented milk products. For the most extensively studied LAB, Lactococcus lactis, a model for casein proteolysis, transport, peptidolysis, and regulation thereof is now established. In addition to nutrient processing, cellular proteolysis plays a critical role in polypeptide quality control and in many regulatory circuits by keeping basal levels of regulatory proteins low and removing them when they are no longer needed. As part of the industrial processes, LAB are challenged by various stress conditions that are likely to affect metabolic activities, including proteolysis. While environmental stress responses of LAB have received increasing interest in recent years, our current knowledge on stress-related proteolysis in LAB is almost exclusively based on studies on L. lactis. This review provides the current status in the research of proteolytic systems of LAB with industrial relevance.
576 citations
TL;DR: Keratinases stand out among proteases since they attack the keratin residues and hence find application in developing cost-effective feather by-products for feed and fertilizers and their prospective application in the challenging field of prion degradation would revolutionize the protease world in the near future.
Abstract: Microbial keratinases have become biotechnologically important since they target the hydrolysis of highly rigid, strongly cross-linked structural polypeptide “keratin” recalcitrant to the commonly known proteolytic enzymes trypsin, pepsin and papain. These enzymes are largely produced in the presence of keratinous substrates in the form of hair, feather, wool, nail, horn etc. during their degradation. The complex mechanism of keratinolysis involves cooperative action of sulfitolytic and proteolytic systems. Keratinases are robust enzymes with a wide temperature and pH activity range and are largely serine or metallo proteases. Sequence homologies of keratinases indicate their relatedness to subtilisin family of serine proteases. They stand out among proteases since they attack the keratin residues and hence find application in developing cost-effective feather by-products for feed and fertilizers. Their application can also be extended to detergent and leather industries where they serve as specialty enzymes. Besides, they also find application in wool and silk cleaning; in the leather industry, better dehairing potential of these enzymes has led to the development of greener hair-saving dehairing technology and personal care products. Further, their prospective application in the challenging field of prion degradation would revolutionize the protease world in the near future.
571 citations
TL;DR: This review discusses the occurrence and structure of chitin, the types and sources of ch itinases, their mode of action, chit inase production, as well as molecular cloning and protein engineering of Chitinases and their biotechnological applications.
Abstract: Chitin and chitinases (EC 3.2.1.14) have an immense potential. Chitinolytic enzymes have wide-ranging applications such as preparation of pharmaceutically important chitooligosaccharides and N-acetyl D-glucosamine, preparation of single-cell protein, isolation of protoplasts from fungi and yeast, control of pathogenic fungi, treatment of chitinous waste, and control of malaria transmission. In this review, we discuss the occurrence and structure of chitin, the types and sources of chitinases, their mode of action, chitinase production, as well as molecular cloning and protein engineering of chitinases and their biotechnological applications.
499 citations
TL;DR: A short overview on the biodegradability and biodegradation of PLA is provided, focused mainly on microbial and enzymatic degradation.
Abstract: Poly(lactide) (PLA) has been developed and made commercially available in recent years. One of the major tasks to be taken before the widespread application of PLA is the fundamental understanding of its biodegradation mechanisms. This paper provides a short overview on the biodegradability and biodegradation of PLA. Emphasis is focused mainly on microbial and enzymatic degradation. Most of the PLA-degrading microorganisms phylogenetically belong to the family of Pseudonocardiaceae and related genera such as Amycolatopsis, Lentzea, Kibdelosporangium, Streptoalloteichus, and Saccharothrix. Several proteinous materials such as silk fibroin, elastin, gelatin, and some peptides and amino acids were found to stimulate the production of enzymes from PLA-degrading microorganisms. In addition to proteinase K from Tritirachium album, subtilisin, a microbial serine protease and some mammalian serine proteases such as alpha-chymotrypsin, trypsin, and elastase could also degrade PLA.
483 citations
TL;DR: PEM surface area was shown to limit power output when the surface area of the PEM was smaller than that of the electrodes due to an increase in internal resistance.
Abstract: Power generation in microbial fuel cells (MFCs) is a function of the surface areas of the proton exchange membrane (PEM) and the cathode relative to that of the anode. To demonstrate this, the sizes of the anode and cathode were varied in two-chambered MFCs having PEMs with three different surface areas (APEM=3.5, 6.2, or 30.6 cm2). For a fixed anode and cathode surface area (AAn=ACat=22.5 cm2), the power density normalized to the anode surface area increased with the PEM size in the order 45 mW/m2 (APEM=3.5 cm2), 68 mW/m2 (APEM=6.2 cm2), and 190 mW/m2 (APEM=30.6 cm2). PEM surface area was shown to limit power output when the surface area of the PEM was smaller than that of the electrodes due to an increase in internal resistance. When the relative cross sections of the PEM, anode, and cathode were scaled according to 2ACat=APEM=2AAn, the maximum power densities of the three different MFCs, based on the surface area of the PEM (APEM=3.5, 6.2, or 30.6 cm2), were the same (168±4.53 mW/m2). Increasing the ionic strength and using ferricyanide at the cathode also increased power output.
464 citations
TL;DR: Traditional methods and methods based on magnetic particles for nucleic acid purification, and the synthesis of a variety of magnetic particles is presented in more detail.
Abstract: Nucleic acid separation is an increasingly important tool for molecular biology. Before modern technologies could be used, nucleic acid separation had been a time- and work-consuming process based on several extraction and centrifugation steps, often limited by small yields and low purities of the separation products, and not suited for automation and up-scaling. During the last few years, specifically functionalised magnetic particles were developed. Together with an appropriate buffer system, they allow for the quick and efficient purification directly after their extraction from crude cell extracts. Centrifugation steps were avoided. In addition, the new approach provided for an easy automation of the entire process and the isolation of nucleic acids from larger sample volumes. This review describes traditional methods and methods based on magnetic particles for nucleic acid purification. The synthesis of a variety of magnetic particles is presented in more detail. Various suppliers of magnetic particles for nucleic acid separation as well as suppliers offering particle-based kits for a variety of different sample materials are listed. Furthermore, commercially available manual magnetic separators and automated systems for magnetic particle handling and liquid handling are mentioned.
450 citations
TL;DR: Partial nitrification to nitrite was reported to be technically feasible and economically favorable, especially when wastewater with high ammonium concentrations or low C/N ratios is treated.
Abstract: Presently, the wastewater treatment practices can be significantly improved through the introduction of new microbial treatment technologies. To meet increasingly stringent discharge standards, new applications and control strategies for the sustainable removal of ammonium from wastewater have to be implemented. Partial nitrification to nitrite was reported to be technically feasible and economically favorable, especially when wastewater with high ammonium concentrations or low C/N ratios is treated. For successful implementation of the technology, the critical point is how to maintain partial nitrification of ammonium to nitrite. Partial nitrification can be obtained by selectively inhibiting nitrite oxidizing bacteria through appropriate regulation of the system's DO concentration, microbial SRT, pH, temperature, substrate concentration and load, operational and aeration pattern, and inhibitor. The review addressed the microbiology, its consequences for their application, the current status regarding application, and the future developments.
433 citations
TL;DR: In vitro studies show the potential of P-solubilizing microorganisms for the simultaneous synthesis and release of pathogen-suppressing metabolites, mainly siderophores, phytohormones, and lytic enzymes, which benefit plant growth and development.
Abstract: Phosphate (P)-solubilizing microorganisms as a group form an important part of the microorganisms, which benefit plant growth and development. Growth promotion and increased uptake of phosphate are not the only mechanisms by which these microorganisms exert a positive effect on plants. Microbially mediated solubilization of insoluble phosphates through release of organic acids is often combined with production of other metabolites, which take part in biological control against soilborne phytopathogens. In vitro studies show the potential of P-solubilizing microorganisms for the simultaneous synthesis and release of pathogen-suppressing metabolites, mainly siderophores, phytohormones, and lytic enzymes. Further trends in this field are discussed, suggesting a number of biotechnological approaches through physiological and biochemical studies using various microorganisms.
TL;DR: Oils and fats are the most important renewable raw materials of the chemical industry making available fatty acids in such purity that they may be used for chemical conversions and for the synthesis of chemically pure compounds.
Abstract: Oils and fats are the most important renewable raw materials of the chemical industry. They make available fatty acids in such purity that they may be used for chemical conversions and for the synthesis of chemically pure compounds. Oleic acid (1) from “new sunflower,” linoleic acid (2) from soybean, linolenic acid (3) from linseed, erucic acid (4) from rape seed, and ricinoleic acid (5) from castor oil are most important for chemical transformations offering in addition to the carboxy group one or more C-C-double bonds. New plant oils containing fatty acids with new and interesting functionalities such as petroselinic acid (6) from Coriandrum sativum, calendic acid (7) from Calendula officinalis, α-eleostearic acid (8) from tung oil, santalbic acid (9) from Santalum album (Linn.), and vernolic acid (10) from Vernonia galamensis are becoming industrially available. The basic oleochemicals are free fatty acids, methyl esters, fatty alcohols, and fatty amines as well as glycerol as a by-product. Their interesting new industrial applications are the usage as environmentally friendly industrial fluids and lubricants, insulating fluid for electric utilities such as transformers and additive to asphalt. Modern methods of synthetic organic chemistry including enzymatic and microbial transformations were applied extensively to fatty compounds for the selective functionalization of the alkyl chain. Syntheses of long-chain diacids, ω-hydroxy fatty acids, and ω-unsaturated fatty acids as base chemicals derived from vegetable oils were developed. Interesting applications were opened by the epoxidation of C-C-double bonds giving the possibility of photochemically initiated cationic curing and access to polyetherpolyols. Enantiomerically pure fatty acids as part of the chiral pool of nature can be used for the synthesis of nonracemic building blocks.
TL;DR: The purpose of this review is to summarise the current state of development of protein separation using magnetic adsorbent particles and identify the obstacles that must be overcome if protein purification with magnetic ads absorbent particles is to find its way into industrial practice.
Abstract: The application of functionalised magnetic adsorbent particles in combination with magnetic separation techniques has received considerable attention in recent years. The magnetically responsive nature of such adsorbent particles permits their selective manipulation and separation in the presence of other suspended solids. Thus, it becomes possible to magnetically separate selected target species directly out of crude biological process liquors (e.g. fermentation broths, cell disruptates, plasma, milk, whey and plant extracts) simply by binding them on magnetic adsorbents before application of a magnetic field. By using magnetic separation in this way, the several stages of sample pretreatment (especially centrifugation, filtration and membrane separation) that are normally necessary to condition an extract before its application on packed bed chromatography columns, may be eliminated. Magnetic separations are fast, gentle, scaleable, easily automated, can achieve separations that would be impossible or impractical to achieve by other techniques, and have demonstrated credibility in a wide range of disciplines, including minerals processing, wastewater treatment, molecular biology, cell sorting and clinical diagnostics. However, despite the highly attractive qualities of magnetic methods on a process scale, with the exception of wastewater treatment, few attempts to scale up magnetic operations in biotechnology have been reported thus far. The purpose of this review is to summarise the current state of development of protein separation using magnetic adsorbent particles and identify the obstacles that must be overcome if protein purification with magnetic adsorbent particles is to find its way into industrial practice.
TL;DR: The experience gained in the Soviet Union forms a promising basis for the development of modern large-scale processes to replace a considerable fraction of the current chemical production of fuel for the authors' future needs on a sustainable basis.
Abstract: Clostridial acetone–butanol fermentation from renewable carbohydrates used to be the largest biotechnological process second only to yeast ethanol fermentation and the largest process ever run under sterile conditions. With the rising prices for mineral oil, it has now the economical and technological potential to replace petrochemistry for the production of fuels from renewable resources. Various methods for using non-food biomass such as cellulose and hemicellulose in agricultural products and wastes have been developed at laboratory scale. To our knowledge, the AB plants in Russia were the only full-scale industrial plants which used hydrolyzates of lignocellosic waste for butanol fermentation. These plants were further developed into the 1980s, and the process was finally run in a continual mode different from plants in Western countries. A biorefinery concept for the use of all by-products has been elaborated and was partially put into practice. The experience gained in the Soviet Union forms a promising basis for the development of modern large-scale processes to replace a considerable fraction of the current chemical production of fuel for our future needs on a sustainable basis.
TL;DR: Eight Escherichia coli strains were studied in minimal medium with a continuous flow system using confocal microscopy and motility affected the biofilm architecture as those strains which had poor motility formed flatter microcolonies compared with those which had enhanced motility.
Abstract: Eight Escherichia coli strains were studied in minimal medium with a continuous flow system using confocal microscopy. K12 wild-type strains ATCC 25404 and MG1655 formed the best biofilms (∼43 μm thick, 21 to 34% surface coverage). JM109, DH5α, and MG1655 motA formed intermediate biofilms (∼13 μm thick, 41 to 58% surface coverage). BW25113, MG1655 qseB, and MG1655 fliA had poor biofilms (surface coverage less than 5%). The best biofilm-formers, ATCC 25404 and MG1655, displayed the highest motility, whereas the worst biofilm former, BW25113, was motility-impaired. The differences in motility were due to differences in expression of the motility loci qseB, flhD, fliA, fliC, and motA (e.g., qseB expression in MG1655 was 139-fold higher than BW25113 and 209-fold higher than JM109). Motility affected the biofilm architecture as those strains which had poor motility (E. coli JM109, E. coli MG1655 motA, and DH5α) formed flatter microcolonies compared with MG1655 and ATCC 25404, which had more dramatic vertical structures as a result of their enhanced motility. The presence of flagella was also found to be important as qseB and fliA mutants (which lack flagella) had less biofilm than the isogenic paralyzed motA strain (threefold less thickness and 15-fold less surface coverage).
TL;DR: E ectoines, as the first extremolytes that are produced in a large scale, have already found application as cell protectants in skin care and as protein-free stabilizers of proteins and cells in life sciences.
Abstract: Extremophilic microorganisms have adopted a variety of ingenious strategies for survival under high or low temperature, extreme pressure, and drastic salt concentrations. A novel application area for extremophiles is the use of “extremolytes,” organic osmolytes from extremophilic microorganisms, to protect biological macromolecules and cells from damage by external stresses. In extremophiles, these low molecular weight compounds are accumulated in response to increased extracellular salt concentrations, but also as a response to other environmental changes, e.g., increased temperature. Extremolytes minimize the denaturation of biopolymers that usually occurs under conditions of water stress and are compatible with the intracellular machinery at high (>1 M) concentrations. The ectoines, as the first extremolytes that are produced in a large scale, have already found application as cell protectants in skin care and as protein-free stabilizers of proteins and cells in life sciences. In addition to ectoines, a range of extremolytes with heterogenous chemical structures like the polyol phosphates di-myoinositol-1,1′-phosphate, cyclic 2,3-diphosphoglycerate, and α-diglycerol phosphate and the mannose derivatives mannosylglycerate (firoin) and mannosylglyceramide (firoin-A) were characterized and were shown to have protective properties toward proteins and cells. A range of new applications, all based on the adaptation to stress conditions conferred by extremolytes, is in development.
TL;DR: This review focuses on the recent concept of molecular understanding of aminoglycoside action and resistance, and describes how enzymatic inactivation through the expression of enzymes, which covalently modify these antibiotics, occurs.
Abstract: Aminoglycosides are potent bactericidal antibiotics targeting the bacterial ribosome, where they bind to the A-site and disrupt protein synthesis. They are particularly active against aerobic, Gram-negative bacteria and act synergistically against certain Gram-positive organisms. Aminoglycosides are used in the treatment of severe infections of the abdomen and urinary tract, bacteremia, and endocarditis. They are also used for prophylaxis, especially against endocarditis. Bacterial resistance to aminoglycosides continues to escalate and is widely recognized as a serious health threat. This might be the reason for the interest in understanding the mechanisms of resistance. It is now clear that the resistance occurs by different mechanisms such as prevention of drug entry, active extrusion of drugs, alteration of the drug target (mutational modification of 16S rRNA and mutational modification of ribosomal proteins), and enzymatic inactivation through the expression of enzymes, which covalently modify these antibiotics. Enzymatic inactivation is normally due to acetyltransferases, nucleotidyltransferases, and phosphotransferases. In this review, we focus on the recent concept of molecular understanding of aminoglycoside action and resistance.
TL;DR: A strong relationship between PPP genes and furfural tolerance is demonstrated and four mutants, zwf1, gnd1, rpe1, and tkl1, which represent genes encoding pentose phosphate pathway (PPP) enzymes are focused on.
Abstract: Engineering yeast to be more tolerant to fermentation inhibitors, furfural and 5-hydroxymethylfurfural (HMF), will lead to more efficient lignocellulose to ethanol bioconversion. To identify target genes involved in furfural tolerance, a Saccharomyces cerevisiae gene disruption library was screened for mutants with growth deficiencies in the presence of furfural. It was hypothesized that overexpression of these genes would provide a growth benefit in the presence of furfural. Sixty two mutants were identified whose corresponding genes function in a wide spectrum of physiological pathways, suggesting that furfural tolerance is a complex process. We focused on four mutants, zwf1, gnd1, rpe1, and tkl1, which represent genes encoding pentose phosphate pathway (PPP) enzymes. At various concentrations of furfural and HMF, a clear association with higher sensitivity to these inhibitors was demonstrated in these mutants. PPP mutants were inefficient at reducing furfural to the less toxic furfuryl alcohol, which we propose is a result of an overall decreased abundance of reducing equivalents or to NADPH's role in stress tolerance. Overexpression of ZWF1 in S. cerevisiae allowed growth at furfural concentrations that are normally toxic. These results demonstrate a strong relationship between PPP genes and furfural tolerance and provide additional putative target genes involved in furfural tolerance.
TL;DR: The biofilm showed a high viability even after 60 days of incubation, apparently due to polyethylene biodegradation, and was found to be up to 2.5 folds higher than that of the protein.
Abstract: We have recently isolated a biofilm-producing strain (C208) of Rhodococcus ruber that degraded polyethylene at a rate of 0.86% per week (r2=0.98). Strain C208 adheres to polyethylene immediately upon exposure to the polyolefin. This initial biofilm differentiates (in a stepwise process that lasts about 20 h) into cell-aggregation-forming microcolonies. Further organization yields "mushroom-like" three-dimensional structures on the mature biofilm. The ratio between the population densities of the biofilm and the planktonic C208 cells after 10 days of incubation was about 60:1, indicating a high preference for the biofilm mode of growth. Analysis of extracellular polymeric substances (EPS) in the biofilm of C208 revealed that the polysaccharides level was up to 2.5 folds higher than that of the protein. The biofilm showed a high viability even after 60 days of incubation, apparently due to polyethylene biodegradation.
TL;DR: The principle of quantitative real-time polymerase chain reaction, its detection reagents, target DNAs, quantification procedures, and affecting factors are reviewed.
Abstract: Due to the advanced development of fluorogenic chemistry, quantitative real-time polymerase chain reaction (qRT-PCR) has become an emerging technique for the detection and quantification of microorganisms in the environment. Compared with the conventional hybridization- and PCR-based techniques, qRT-PCR not only has better sensitivity and reproducibility, but it is also quicker to perform and has a minimum risk of amplicon carryover contamination. This article reviews the principle of this emerging technique, its detection reagents, target DNAs, quantification procedures, and affecting factors. The applications of qRT-PCR for the quantification of microorganisms in the environment are also summarized.
TL;DR: An isolation program targeting Thraustochytrids (marine fungoid protists) from 19 different Atlantic Canadian locations was performed, and this strain, ONC-T18, was found to produce the carotenoids and xanthophylls astaxanthin, zeaxanth in, canthaxanthIn, echinenone, and β-carotene.
Abstract: An isolation program targeting Thraustochytrids (marine fungoid protists) from 19 different Atlantic Canadian locations was performed. Sixty-eight isolates were screened for biomass, total fatty acid (TFA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) content. Analysis of fatty acid methyl ester results discerned four distinctive clusters based on fatty acid profiles, with biomass ranging from 0.1 to 2.3 g L(-1), and lipid, EPA, and DHA contents ranging from 27.1 to 321.14, 2.97 to 21.25, and 5.18 to 83.63 mg g(-1) biomass, respectively. ONC-T18, was subsequently chosen for further manipulations. Identified using 18S rRNA gene sequencing techniques as a Thraustochytrium sp., most closely related to Thraustochytrium striatum T91-6, ONC-T18 produced up to 28.0 g L(-1) biomass, 81.7% TFA, 31.4% (w/w biomass) DHA, and 4.6 g L(-1) DHA under optimal fermentation conditions. Furthermore, this strain was found to produce the carotenoids and xanthophylls astaxanthin, zeaxanthin, canthaxanthin, echinenone, and beta-carotene. Given this strain's impressive productivity when compared to commercial strains, such as Schizochytrium sp. SR21 (which has only 50% TFA), coupled with its ability to grow at economical nitrogen and very low salt concentrations (2 g L(-1)), ONC-T18 is seen as an ideal candidate for both scale-up and commercialization.
TL;DR: In this review, advances in metabolic engineering for the production of the aromatic amino acids and useful aromatic intermediates are described with particular emphasis on two representative producer organisms, Corynebacterium glutamicum and Escherichia coli.
Abstract: The aromatic amino acids, L-tryptophan, L-phenylalanine, and L-tyrosine, can be manufactured by bacterial fermentation. Until recently, production efficiency of classical aromatic amino-acid-producing mutants had not yet reached a high level enough to make the fermentation method the most economic. With the introduction of recombinant DNA technology, it has become possible to apply more rational approaches to strain improvement. Many recent activities in this metabolic engineering have led to several effective approaches, which include modification of terminal pathways leading to removal of bottleneck or metabolic conversion, engineering of central carbon metabolism leading to increased supply of precursors, and transport engineering leading to reduced intracellular pool of the aromatic amino acids. In this review, advances in metabolic engineering for the production of the aromatic amino acids and useful aromatic intermediates are described with particular emphasis on two representative producer organisms, Corynebacterium glutamicum and Escherichia coli.
TL;DR: How the active site topologies of these glycoside hydrolases influence their mode of action in heterogeneous phase is examined, to discuss the next challenges in the basic and applied field of the galactans of red algae and of their related degrading microorganisms.
Abstract: Agars and carrageenans are 1,3-alpha-1,4-beta-galactans from the cell walls of red algae, substituted by zero (agarose), one (kappa-), two (iota-), or three (lambda-carrageenan) sulfate groups per disaccharidic monomer. Agars, kappa-, and iota-carrageenans auto-associate into crystalline fibers and are well known for their gelling properties, used in a variety of laboratory and industrial applications. These sulfated galactans constitute a crucial carbon source for a number of marine bacteria. These microorganisms secrete glycoside hydrolases specific for these polyanionic, insoluble polysaccharides, agarases and carrageenases. This article reviews the microorganisms involved in the degradation of agars and carrageenans, in their environmental and taxonomic diversity. We also present an overview on the biochemistry of the different families of galactanases. The structure-function relationships of the family GH16 beta-agarases and kappa-caraggeenases and of the family GH82 iota-carrageenases are discussed in more details. In particular, we examine how the active site topologies of these glycoside hydrolases influence their mode of action in heterogeneous phase. Finally, we discuss the next challenges in the basic and applied field of the galactans of red algae and of their related degrading microorganisms.
TL;DR: Results demonstrate that the fermentation of the lignocellulose hydrolysate is performed efficiently by the recombinant Saccharomyces strain with abilities for xylose assimilation and cellooligosaccharide degradation.
Abstract: The sulfuric acid hydrolysate of lignocellulosic biomass, such as wood chips, from the forest industry is an important material for fuel bioethanol production. In this study, we constructed a recombinant yeast strain that can ferment xylose and cellooligosaccharides by integrating genes for the intercellular expressions of xylose reductase and xylitol dehydrogenase from Pichia stipitis, and xylulokinase from Saccharomyces cerevisiae and a gene for displaying β-glucosidase from Aspergillus acleatus on the cell surface. In the fermentation of the sulfuric acid hydrolysate of wood chips, xylose and cellooligosaccharides were completely fermented after 36 h by the recombinant strain, and then about 30 g/l ethanol was produced from 73 g/l total sugar added at the beginning. In this case, the ethanol yield of this recombinant yeast was much higher than that of the control yeast. These results demonstrate that the fermentation of the lignocellulose hydrolysate is performed efficiently by the recombinant Saccharomyces strain with abilities for xylose assimilation and cellooligosaccharide degradation.
TL;DR: A second fungal haloperoxidase of the heme-thiolate type has been discovered in the agaric mushroom Agrocybe aegerita and could become promising biocatalysts in biotechnological applications because they seemingly fill the gap between CPO and P450 enzymes and act as “self-sufficient” peroxygenases.
Abstract: Heme-thiolate haloperoxidases are undoubtedly the most versatile biocatalysts of the hemeprotein family and share catalytic properties with at least three further classes of heme-containing oxidoreductases, namely, classic plant and fungal peroxidases, cytochrome P450 monooxygenases, and catalases. For a long time, only one enzyme of this type--the chloroperoxidase (CPO) of the ascomycete Caldariomyces fumago--has been known. The enzyme is commercially available as a fine chemical and catalyzes the unspecific chlorination, bromination, and iodation (but no fluorination) of a variety of electrophilic organic substrates via hypohalous acid as actual halogenating agent. In the absence of halide, CPO resembles cytochrome P450s and epoxidizes and hydroxylates activated substrates such as organic sulfides and olefins; aromatic rings, however, are not susceptible to CPO-catalyzed oxygen-transfer. Recently, a second fungal haloperoxidase of the heme-thiolate type has been discovered in the agaric mushroom Agrocybe aegerita. The UV-Vis adsorption spectrum of the isolated enzyme shows little similarity to that of CPO but is almost identical to a resting-state P450. The Agrocybe aegerita peroxidase (AaP) has strong brominating as well as weak chlorinating and iodating activities, and catalyzes both benzylic and aromatic hydroxylations (e.g., of toluene and naphthalene). AaP and related fungal peroxidases could become promising biocatalysts in biotechnological applications because they seemingly fill the gap between CPO and P450 enzymes and act as "self-sufficient" peroxygenases. From the environmental point of view, the existence of a halogenating mushroom enzyme is interesting because it could be linked to the multitude of halogenated compounds known from these organisms.
TL;DR: The nucleic acid analogs provide an advantage to design and implementation, therapies, and research assays, which were not implemented due to limitations associated with standard nucleic acids chemistry.
Abstract: Nucleic acid analogs and mimics are commonly the modifications of native nucleic acids at the nucleobase, the sugar ring, or the phosphodiester backbone Many forms of promising nucleic acid analogs and mimics are available, such as locked nucleic acids (LNAs), peptide nucleic acids (PNAs), and morpholinos LNAs, PNAs, and morpholinos can form both duplexes and triplexes and have improved biostability They have become a general and versatile tool for DNA and RNA recognition LNA is a general and versatile tool for specific, high-affinity recognition of single-stranded DNA (ssDNA) and single-stranded RNA (ssRNA) LNA can be used for designing LNA oligoes for hybridization studies or as real time polymerase chain reaction probes in the form of Taqman probes LNA also has therapeutic and diagnostic applications PNA is another type of DNA analog with neutral charge The extreme stability of PNA makes it an ideal candidate for the antisense and antigene application PNA is used as probe for gene cloning, mutation detection, and in homologous recombination studies It was also used to design transcription factor decoy molecules for target gene induction Morpholino, another structural type, was devised to circumvent cost problems associated with DNA analogs It has become the premier knockdown tool in developmental biology due to its cytosolic delivery in the embryos by microinjection Thus, the nucleic acid analogs provide an advantage to design and implementation, therapies, and research assays, which were not implemented due to limitations associated with standard nucleic acids chemistry
TL;DR: Screening phage display libraries of proteins and peptides has, for almost two decades, proven to be a powerful technology for selecting polypeptides with desired biological and physicochemical properties from huge molecular libraries.
Abstract: Screening phage display libraries of proteins and peptides has, for almost two decades, proven to be a powerful technology for selecting polypeptides with desired biological and physicochemical properties from huge molecular libraries. The scope of phage display applications continues to expand. Recent applications and technical improvements driving further developments in the field of phage display are discussed.
TL;DR: A brief survey of research performed in the production of poly(hydroxyalkanoate) polymers, with specific emphasis on studies describing the utilization of intact triacylglycerols, dairy whey, molasses, and meat-and-bone meal as substrates in the microbial synthesis of PHA polymers.
Abstract: Aside from their importance to the survival and general welfare of mankind, agriculture and its related industries produce large quantities of feedstocks and coproducts that can be used as inexpensive substrates for fermentative processes. Successful adoption of these materials into commercial processes could further the realization of a biorefinery industry based on agriculturally derived feedstocks. One potential concept is the production of poly(hydroxyalkanoate) (PHA) polymers, a family of microbial biopolyesters with a myriad of possible monomeric compositions and performance properties. The economics for the fermentative production of PHA could benefit from the use of low-cost agricultural feedstocks and coproducts. This mini-review provides a brief survey of research performed in this area, with specific emphasis on studies describing the utilization of intact triacylglycerols (vegetable oils and animal fats), dairy whey, molasses, and meat-and-bone meal as substrates in the microbial synthesis of PHA polymers.
TL;DR: This review considers the motivations for bioreporter developments and discusses the suitability of extant bioreporters for the proposed applications to stimulate complementary research and to help researchers to develop realistic objectives.
Abstract: Since the development of the first whole-cell living biosensor or bioreporter about 15 years ago, construction and testing of new genetically modified microorganisms for environmental sensing and reporting has proceeded at an ever increasing rate. One and a half decades appear as a reasonable time span for a new technology to reach the maturity needed for application and commercial success. It seems, however, that the research into cellular biosensors is still mostly in a proof-of-principle or demonstration phase and not close to extensive or commercial use outside of academia. In this review, we consider the motivations for bioreporter developments and discuss the suitability of extant bioreporters for the proposed applications to stimulate complementary research and to help researchers to develop realistic objectives. This includes the identification of some popular misconceptions about the qualities and shortcomings of bioreporters.
TL;DR: Heterofermentative lactic acid bacteria such as Leuconostoc, Oenococcus, and Lactobacillus strains are able to use a variety of unusual fermentation reactions in addition to classical heterofermentation, which are significant for food biotechnology/microbiology.
Abstract: Heterofermentative lactic acid bacteria (LAB) such as Leuconostoc, Oenococcus, and Lactobacillus strains ferment pentoses by the phosphoketolase pathway. The extra NAD(P)H, which is produced during growth on hexoses, is transferred to acetyl-CoA, yielding ethanol. Ethanol fermentation represents the limiting step in hexose fermentation, therefore, part of the extra NAD(P)H is used to produce erythritol and glycerol. Fructose, pyruvate, citrate, and O2 can be used in addition as external electron acceptors for NAD(P)H reoxidation. Use of the external acceptors increases the growth rate of the bacteria. The bacteria are also able to ferment organic acids like malate, pyruvate, and citrate. Malolactic fermentation generates a proton potential by substrate transport. Pyruvate fermentation sustains growth by pyruvate disproportionation involving pyruvate dehydrogenase. Citrate is fermented in the presence of an additional electron donor to acetate and lactate. Thus, heterofermentative LAB are able to use a variety of unusual fermentation reactions in addition to classical heterofermentation. Most of the reactions are significant for food biotechnology/microbiology.