Optimization of culture conditions and scale-up to plant scales for teicoplanin production by <Emphasis Type="Italic">Actinoplanes teichomyceticus</Emphasis>

This report describes the optimization of culture conditions for teicoplanin production by Actinoplanes teichomyceticus KCCM-10601, an identified high-teicoplanin-producing strain (US 2006/0134757 A1). Among the conditions tested, temperature, pH, and the dissolved oxygen tension (DOT) were key factors affecting teicoplanin production. When the temperature, pH, and DOT were controlled at 34 degrees C, 7.0 and 20-30%, respectively, a dry-cell weight of 42.8 g l(-1) and a teicoplanin production of 2.9 g l(-1) were obtained after 120 h of batch culture, corresponding to a specific teicoplanin content of 67.8 mg g-DCW(-1). Teicoplanin production was scaled-up from a laboratory scale (7-l fermenter) to a pilot scale (300 l) and a plant scale (5,000 l) using the impeller tip velocity (V tip) as a scale-up parameter. Teicoplanin production at the laboratory scale was similar to those at the pilot and plant scales. This is the highest report of pilot- and plant-scale production of teicoplanin.     

Biosynthesis and biotechnological production of flavanones: current state and perspectives

Polyphenols produced in a wide variety of flowering and fruit-bearing plants have the potential to be valuable fine chemicals for the treatment of an assortment of human maladies. One of the major constituents within this chemical class are flavonoids, among which flavanones, as the precursor to all flavonoid structures, are the most prevalent. We review the current status of flavanone production technology using microorganisms, with focus on heterologous protein expression. Such processes appear as attractive production alternatives for commercial synthesis of these high-value chemicals as traditional chemical, and plant cell cultures have significant drawbacks. Other issues of importance, including fermentation configurations and economics, are also considered.     

Biosynthesis and production of sabinene: current state and perspectives

Applied Microbiology and Biotechnology - Sabinene is an important naturally occurring bicyclic monoterpene which can be used as flavorings, perfume additives, fine chemicals, and advanced biofuels....     

Artemisinin: current state and perspectives for biotechnological production of an antimalarial drug

Artemisinin isolated from the aerial parts of Artemisia annua L. is a promising and potent antimalarial drug which has a remarkable activity against chloroquine-resistant and chloroquine-sensitive strains of Plasmodium falciparum, and is useful in treatment of cerebral malaria. Because the low content (0.01–1 %) of artemisinin in A. annua is a limitation to the commercial production of the drug, many research groups have been focusing their researches on enhancing the production of artemisinin in tissue culture or in the whole plant of A. annua. This review mainly focuses on the progresses made in the production of artemisinin from A. annua by biotechnological strategies including in vitro tissue culture, metabolic regulation of artemisinin biosynthesis, genetic engineering, and bioreactor technology.     

Ethanol production from xylose in engineered Saccharomyces cerevisiae strains: current state and perspectives

Bioethanol production from xylose is important for utilization of lignocellulosic biomass as raw materials. The research on yeast conversion of xylose to ethanol has been intensively studied especially for genetically engineered Saccharomyces cerevisiae during the last 20 years. S. cerevisiae, which is a very safe microorganism that plays a traditional and major role in industrial bioethanol production, has several advantages due to its high ethanol productivity, as well as its high ethanol and inhibitor tolerance. However, this yeast cannot ferment xylose, which is the dominant pentose sugar in hydrolysates of lignocellulosic biomass. A number of different strategies have been applied to engineer yeasts capable of efficiently producing ethanol from xylose, including the introduction of initial xylose metabolism and xylose transport, changing the intracellular redox balance, and overexpression of xylulokinase and pentose phosphate pathways. In this review, recent progress with regard to these studies is discussed, focusing particularly on xylose-fermenting strains of S. cerevisiae. Recent studies using several promising approaches such as host strain selection and adaptation to obtain further improved xylose-utilizing S. cerevisiae are also addressed.     

Application of conjugation using ϕC31 <Emphasis Type="Italic">att/int</Emphasis> system for <Emphasis Type="Italic">Actinoplanes teichomyceticus</Emphasis>, a producer of teicoplanin

Actinoplanes teichomyceticus produces teicoplanin, which is a glycopeptide antibiotic for Gram-positive pathogenic bacteria and methicillin-resistant Staphylococcus aureus (MRSA). For a molecular genetic study of A. teichomyceticus, an effective transformation method using the conjugal transfer of DNA from E. coli to spores of A. teichomyceticus was established for the first time, based on the bacteriophage ϕC31 att/int system, in the genus of Actinoplanes. The high frequency of transconjugation was obtained on MS medium containing 40 mM MgCl₂, using 1.25 × 10⁸ E. coli donor cells and 10⁵ spores without a heat treatment. In addition, by cloning and sequencing the attB site A. teichomyceticus was shown to contain a single attB site within an ORF coding for a pirin homolog. Also, its attB site sequence showed high homology to that of Streptomyces lividans, unlike the case of Kitasatospora setae despite being a non-Streptomyces actinomycete, which seems to be closely related to the high transconjugation frequency of A. teichomyceticus.     

Production of teicoplanin by a mutant of Actinoplanes teicomyceticus

Teicoplanin, a glucopeptide antibiotic, was produced by a mutant of Actinoplanes teicomyceticus at 300 mg l^–1 using mannose and yeast extract as carbon and nitrogen sources in flask culture and at 500 mg l^–1 in 5-l jar fermenter. Teicoplanin production was 25-fold higher than in the parent strain.     

Microbiological production of tocopherols: current state and prospects

Tocopherols are antioxidants that prevent various diseases caused by oxidative stress. Tocochromanols comprise four isoforms of tocopherols and four isoforms of tocotrienols but α-tocopherol is the most abundant and active isoform in human and animal tissues. Tocopherols are used as dietary supplements for human, as food preservatives, in manufacture of cosmetics, and for fortification of animal feed. Only photosynthetic cells are known to accumulate detectable concentrations of tocopherols. Tocopherols can be extracted and purified or concentrated from vegetable oils and other higher plant materials. However, the concentrations in these higher plant materials are very low and there are high proportions of the less-active homologues of tocopherols. Among the many strains of photosynthetic microorganisms known to accumulate tocopherols, Euglena gracilis is promising for commercial production of α-tocopherol. The growth rate and α-tocopherol contents are relatively high and α-tocopherol comprise more than 97% of all the tocopherols accumulated by Euglena gracilis. Although a lot of work has been done to increase the contents and composition of tocopherols in higher plants through genetic and metabolic engineering, work on genetic modification of microorganisms for increased tocopherol accumulation is scarce. Many cultivation systems have been investigated for efficient production of tocopherol by Euglena gracilis. However, those that involve heterotrophic metabolism are more promising. Bubble columns and flat-plate photobioreactors are more suitable for commercial production of tocopherols, than the tubular, internally illuminated, and open-air photobioreactors.     

Biogas production: current state and perspectives

Anaerobic digestion of energy crops, residues, and wastes is of increasing interest in order to reduce the greenhouse gas emissions and to facilitate a sustainable development of energy supply. Production of biogas provides a versatile carrier of renewable energy, as methane can be used for replacement of fossil fuels in both heat and power generation and as a vehicle fuel. For biogas production, various process types are applied which can be classified in wet and dry fermentation systems. Most often applied are wet digester systems using vertical stirred tank digester with different stirrer types dependent on the origin of the feedstock. Biogas is mainly utilized in engine-based combined heat and power plants, whereas microgas turbines and fuel cells are expensive alternatives which need further development work for reducing the costs and increasing their reliability. Gas upgrading and utilization as renewable vehicle fuel or injection into the natural gas grid is of increasing interest because the gas can be used in a more efficient way. The digestate from anaerobic fermentation is a valuable fertilizer due to the increased availability of nitrogen and the better short-term fertilization effect. Anaerobic treatment minimizes the survival of pathogens which is important for using the digested residue as fertilizer. This paper reviews the current state and perspectives of biogas production, including the biochemical parameters and feedstocks which influence the efficiency and reliability of the microbial conversion and gas yield.     

Properties and biotechnological applications of halohydrin dehalogenases: current state and future perspectives

Halohydrin dehalogenases (HHDHs) are lyases that catalyze the cleavage of carbon–halogen bond of halohydrins. They also can catalyze the reverse reaction in the presence of nucleophiles such as cyanide, azide, and nitrite ions. HHDHs have been recognized as the ideal tools for the degradation of various halogenated environmental pollutants. Moreover, they can be used as biocatalysts for the kinetic resolution of halohydrins and epoxides, and for the preparation of various substituted alcohols. This review is mainly focused on the current status of research on HHDHs, highlighting the production, characterization, structures and mechanism, protein engineering, and biotechnological applications of HHDHs.     

梭菌属分类研究进展:现状和展望

目前构成梭菌属的微生物在系统发育和表型特征上不一致。多相分类数据表明,梭菌属物种之间差异大。大量基于16S rRNA 基因的系统发育研究表明,梭菌属应被限定为梭菌属类群I,作为狭义梭菌属(Clostridium sensu stricto)。尽管有这方面认识,梭菌属新物种仍持续增加,这些新物种并不能与梭菌属类群I 和标准种丁酸梭菌(C. butryicum)形成一致分支,引发梭菌属分类上的混乱。本文明确了梭菌属物种的范围,即只包括模式种和梭菌属类群I。此外,4 个物种念珠状真杆菌(Eubacterium moniliforme)、旋舞真杆菌(Eubacterium tarantellae)、最大八叠球菌(Sarcina maxima)和胃八叠球菌(Sarcina ventriculi)应被调至梭菌属,分别命名为念珠状梭菌(Clostridium moniliforme)、旋舞梭菌(Clostridium tarantellae)、最大梭菌(Clostridium maximum)和胃梭菌(Clostridium ventriculi)。一个新属哈撒韦氏菌属(Hathewaya)被提议成立,3 个梭菌属物种溶组织梭菌(Clostridium histolyticum)、泥渣梭菌(Clostridium limosum)和解朊梭菌(Clostridiumproteolyticum)重新归为溶组织哈撒韦氏菌(Hathewaya histolytica)、泥渣哈撒韦氏菌(Hathewaya limosa)和解朊哈撒韦氏菌(Hathewaya proteolytica),其中Hathewaya histolytica 为模式种。     

Biotechnological production of eleutherosides: current state and perspectives

Eleutherosides, the phenylpropanoid and lignan glycosides, are the active ingredients accumulated in the roots and stems of Eleutherococcus species and in Eleutherococcus senticosus in particular. Syringin (=eleutheroside B) and (?) syringaresinol-di-O-β-d-glucoside (=eleutheroside E) appear as the most important bioactive compounds which are used as adaptogens, besides their abundant antidiabetic and anticancer properties. As the availability of “Eleuthero” is becoming increasingly limited because of its scanty natural distribution, the production of these compounds by biotechnological means has become an attractive alternative. In E. senticosus and other closely related species, Eleutherococcus sessiliflorus, Eleutherococcus chiisanensis, and Eleutherococcus koreanum, organogenic cultures have been induced for the production of eleutherosides. Bioreactor cultures have been established and various parameters, which influence on the accumulation of biomass and secondary metabolites, have been thoroughly investigated. Pilot-scale cultures have also been accomplished for the large-scale production of somatic embryos containing abundant amounts of eleutherosides. This review describes the biotechnological approaches and challenges for the production of eleutherosides.     

Occurrence, production, and applications of gellan: current state and perspectives

Bacterial exopolysaccharides (EPS) are products of biotechnology that are of high interest due to their rheological properties. This is the case of sphingans, a group of structurally related EPS secreted by members of the genus Sphingomonas. Among these, gellan is a multifunctional gelling agent produced in high yields by the non-pathogenic strain Sphingomonas elodea ATCC 31461. In its native form, gellan is a linear anionic EPS based on a tetrasaccharide repeat unit composed of two molecules of D: -glucose, one of L: -rhamnose and one of D: -glucuronic acid. The native gellan is partially esterified with acyl substituents (1 mol of glycerate and 0.5 mol of acetate) per repeat unit. Gellan has unique characteristics and has many applications, particularly in the food, pharmaceutical, and biomedical fields. This review summarizes current knowledge on the structure and properties of gellan and provides details about the biosynthesis of this exopolysaccharide. In addition, a highlight of the importance of gellan in industrial and medicinal applications is given.     

The lipases from Yarrowia lipolytica: genetics, production, regulation, biochemical characterization and biotechnological applications

Lipases are serine hydrolases that catalyze in nature the hydrolysis of ester bonds of long chain triacylglycerol into fatty acid and glycerol. However, in favorable thermodynamic conditions, they are also able to catalyze reactions of synthesis such as esterification or amidation. The non-conventional yeast Yarrowia lipolytica possesses 16 paralogs of genes coding for lipase. However, little information on all those paralogs has been yet obtained and only three isoenzymes, namely Lip2p, Lip7p and Lip8p have been partly characterized so far. Microarray data suggest that only a few of them could be expressed and that lipase synthesis seems to be dependent on the fatty acid or oil used as carbon source confirming the high adaptation of Y. lipolytica to hydrophobic substrate utilization. This review focuses on the biochemical characterization of those enzymes with special emphasis on the Lip2p lipase which is the isoenzyme mainly synthesized by Y. lipolytica. Crystallographic data highlight that this latter is a lipase sensu stricto with a lid covering the active site of the enzyme in its closed conformation. Recent findings on enzyme conditioning in dehydrated or liquid formulation, in enzyme immobilization by entrapment in natural polymers from either organic or mineral origins are also discussed together with long-term storage strategies. The development of various biotechnological applications in different fields such as cheese ripening, waste treatment, drug synthesis or human therapeutics is also presented.     

Thermophilic,lignocellulolytic bacteria for ethanol production: current state and perspectives

Lignocellulosic biomass contains a variety of carbohydrates, and their conversion into ethanol by fermentation requires an efficient microbial platform to achieve high yield, productivity, and final titer of ethanol. In recent years, growing attention has been devoted to the development of cellulolytic and saccharolytic thermophilic bacteria for lignocellulosic ethanol production because of their unique properties. First of all, thermophilic bacteria possess unique cellulolytic and hemicellulolytic systems and are considered as potential sources of highly active and thermostable enzymes for efficient biomass hydrolysis. Secondly, thermophilic bacteria ferment a broad range of carbohydrates into ethanol, and some of them display potential for ethanologenic fermentation at high yield. Thirdly, the establishment of the genetic tools for thermophilic bacteria has allowed metabolic engineering, in particular with emphasis on improving ethanol yield, and this facilitates their employment for ethanol production. Finally, different processes for second-generation ethanol production based on thermophilic bacteria have been proposed with the aim to achieve cost-competitive processes. However, thermophilic bacteria exhibit an inherent low tolerance to ethanol and inhibitors in the pretreated biomass, and this is at present the greatest barrier to their industrial application. Further improvement of the properties of thermophilic bacteria, together with the optimization production processes, is equally important for achieving a realistic industrial ethanol production.     

Metabolic engineering of strains of Ralstonia eutropha and Pseudomonas putida for biotechnological production of 2-methylcitric acid

In this study strains of Ralstonia eutropha H16 and Pseudomonas putida KT2440 were engineered which are suitable for biotechnological production of 2-methylcitric acid (2MC). Analysis of a previous mutant of R. eutropha able to accumulate 2MC recommended this strain as a candidate for fermentative production of 2MC. This knowledge was used for construction of strains of R. eutropha H16 and P. putida KT2440 capable of enhanced production of 2MC. In both bacteria the chromosomal genes encoding the 2-methyl-cis-aconitate hydratase (acnM) were disrupted by directed insertion of a copy of an additional 2-methylcitrate synthase gene (prpC) yielding strains R. eutropha DeltaacnM(Re)OmegaKmprpC(Pp) and P. putida DeltaacnM(Pp)OmegaKmprpC(Re). In both strains 2-methylcitrate synthase was expressed under control of the constitutive kanamycin-resistance gene (OmegaKm) resulting in up to 20-fold higher specific 2-methylcitrate synthase activities in comparison to the wild type. The disruption of the acnM gene by insertion of prpC led to a propionate- and levulinate-negative phenotype of the engineered strains, and analysis of supernatant of these strains revealed overproduction and accumulation of 2MC in the medium. A two stage cultivation regime comprising an exponential growth phase and a 2MC production phase was developed and applied to both engineered strains for optimum production of 2MC. Whereas gluconate, fructose or succinate were provided as carbon source for the exponential growth phase, a combination of propionate or levulinate as precursor substrate for provision of propionyl-CoA and succinate or fumarate as precursor substrate for provision of oxaloacetate were used in the production phase to make sure that the 2-methylcitrate synthase was provided with their substrates. Employing the optimised feeding regime P. putida DeltaacnM(Pp)OmegaKmprpC(Re) and R. eutropha DeltaacnM(Re)OmegaKmprpC(Pp) produced 2MC up to maximal concentrations of 7.2 g/L or 26.5 mM and 19.2 g/L or 70.5 mM, respectively, during 144 h of cultivation.     

Helicobacter pylori chromosomal DNA replication: current status and future perspectives

Helicobacter pylori causes gastritis, gastric ulcer and gastric cancer. Though DNA replication and its control are central to bacterial proliferation, pathogenesis, virulence and/or dormancy, our knowledge of DNA synthesis in slow growing pathogenic bacteria like H. pylori is still preliminary. Here, we review the current understanding of DNA replication, replication restart and recombinational repair in H. pylori. Several differences have been identified between the H. pylori and Escherichia coli replication machineries including the absence of DnaC, the helicase loader usually conserved in gram-negative bacteria. These differences suggest different mechanisms of DNA replication at initiation and restart of stalled forks in H. pylori.