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Enzyme Production (enzyme + production)
Selected AbstractsEvaluation of a novel Bacillus strain from a north-western Spain hot spring as a source of extracellular thermostable lipaseJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 10 2009Francisco J. Deive Abstract BACKGROUND: Thermophilic microorganisms are receiving significant attention as a source of useful thermostable enzymes. However, the number of known strains is still limited, and very often their most interesting biocatalysts are intracellular or membrane-bound and produced at low levels. Thus, the isolation and study of novel extracellular enzyme-producing thermophilic microorganisms is very interesting. Moreover, the assessment of bioreactor performance is crucial, given the scarce information on the large-scale culture of these strains. RESULTS: The production of a thermostable extracellular lipase in submerged cultures of a thermophilic microorganism, recently isolated in north-west Spain, was investigated. The strain was identified by 16S rDNA sequencing as belonging to genus Bacillus. The influence of operating variables (i.e. pH, temperature, aeration) on lipase biosynthesis was analysed. Enzyme production at bioreactor scale was investigated, special attention being paid to the effect of aeration and agitation rates. CONCLUSION: The best conditions for the studied process were determined in shake flasks as pH 7.0, 55 °C and high aeration levels. Also, the non-association between lipase production and cell growth was ascertained. The culture of this novel strain was successfully carried out in laboratory-scale bioreactors, thus proving its potential for further applications. Copyright © 2009 Society of Chemical Industry [source] Purification and Characterization of Thermostable ,-Amylase from a Newly Isolated Thermophilic Bacillus stearothermophilus GRE 1ASIA-PACIFIC JOURNAL OF CHEMICAL ENGINEERING, Issue 5-6 2005S.M. Zakir Hossain A thermophilic bacterium, Bacillus stearothermophilus GRE 1, isolated from an Ethiopian hyperthermal spring produced a thermostable ,-amylase. Enzyme production in shake flask experiments while using optimum nutrient supplements and environmental conditions was 2.35 U/ml. Under optimized conditions in a bioreactor, 5-6 fold higher enzyme activity was obtained than that of a shake flask. Gel filtration chromatography yielded a purification factor of 33.62-fold and a recovery of 46.52%. The optimum temperature for activity was determined to be 60-70d,C and optimum pH was in the range of 5.5-6.0. The enzyme maintained 50% of its original activity after 45 minutes of incubation at 80d,C, and is stable at pH values of 5.0-9.0. Enzyme activity was strongly inhibited by Cu2+, Zn2+ and Fe2+. The enzyme is calcium independent and 94% and 86% relative activities were displayed with low concentrations of Co2+ and Mg2+, respectively. [source] White-rot fungi combined with lignite granules and lignitic xylite to decolorize textile industry wastewaterENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 1 2010Ulrike Böhmer Abstract The feasibility of using immobilized fungi to decolorize textile industry wastewater containing dyes was examined in experiments with: two species of white-rot fungi (a Marasmius species from Indonesia, which produces copious biomass, and Trametes hirsuta, which produces high levels of laccase); two types of lignite products as adsorbents and solid substrates (lignitic xylite and lignite granules); and four simulated wastewaters, each containing a different kinds of reactive textile azo dye. The growth, extracellular enzyme production, dye degradation and dye absorption parameters afforded by each permutation of fungus, substrate and dye were then measured. Both fungal species grew poorly on xylite, but much better on lignite granules. Marasmius sp. produced up to 67,U/L laccase on lignite granules, but just 10,U/L on xylite, and no other detectable extracellular enzymes. T. hirsuta produced 1343,U/L laccase and up to 12,U/L unspecific peroxidase when immobilized on lignite granules, and 898,U/L laccase with 14,U/L unspecific peroxidase when immobilized on xylite. The amount of color lost from the dye solutions depended on both the type of dye and the enzyme levels in the fermenter. [source] Production of a Laccase and Decrease of the Phenolic Content in Canola Meal during the Growth of the Fungus Pleurotus ostreatus in Solid State Fermentation ProcessesENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 1 2004J. Hu Abstract Solid state fermentation of canola meal was carried out with the fungus Pleurotus ostreatus DAOM 197961, which is a producer of laccase. The aim of this study was to examine the effects of moisture content, inoculum size, homogenisation of inoculum and particle size of canola meal on the growth of the fungus, the production of a laccase and the decrease of the content of sinapic acid esters (SAE) in a solid state process. The results showed that the optimum moisture content, which was varied in the media between 50% and 75%, for the growth and enzyme production was 60%. The initial rate of SAE content decrease was faster in the media with 70% and 75% moisture than in those with lower moisture levels. In the study of the effects of inoculum concentration in the range of 1.1 mg to 5.5 mg/g of the medium, it was found that larger amounts of biomass and enzyme were produced in the media with inoculum concentrations from 1.1 mg to 3.3 mg/g of the medium than in the media with a higher inoculum concentration. The final and approximately the same concentrations of SAE were reached at the same time regardless of the inoculum concentration. Considering that the fungus formed pellets under the conditions at which it was grown during the inoculum preparation, it was necessary to break them by homogenisation prior to their utilisation as an inoculum. The homogenisation was carried out during a period between 15s and 200s. Although higher biomass concentrations and enzyme activities were obtained in the media which were inoculated with the inoculum homogenised for 15s and 30s, the maximum enzyme activities and biomass concentrations were reached in the media inoculated with the inoculum, which was homogenised for 120s and 200s. The time of inoculum homogenisation did not influence the kinetics of the SAE decrease. When the effects of the particle size of canola meal on the process were studied, it was found that larger particles of the meal in the solid media were more favourable for the production of the biomass and enzyme, and for a faster decrease of the SAE content than those of smaller sizes. From the obtained results it can be concluded that the tested variables have a significant influence on the growth of the fungus Pleurotus ostreatus DAOM 197961, the production of laccase and the decrease of the SAE content in canola meal. The data could be useful for the development of a solid state process for the production of laccase and for the decrease of the phenolics content in canola meal. [source] Use of monoclonal antibodies to quantify the dynamics of ,-galactosidase and endo-1,4-,-glucanase production by Trichoderma hamatum during saprotrophic growth and sporulation in peatENVIRONMENTAL MICROBIOLOGY, Issue 5 2005Christopher R. Thornton Summary Trichoderma species are ubiquitous soil and peat-borne saprotrophs that have received enormous scientific interest as biocontrol agents of plant diseases caused by destructive root pathogens. Mechanisms of biocontrol such as antibiosis and hyperparasitism are well documented and the biochemistry and molecular genetics of these processes defined. An aspect of biocontrol that has received little attention is the ability of Trichoderma species to compete for nutrients in their natural environments. Trichoderma species are efficient producers of polysaccharide-degrading enzymes that enable them to colonize organic matter thereby preventing the saprotrophic spread of plant pathogens. This study details the use of monoclonal antibodies (mAbs) to quantify the production of two enzymes implicated in the saprotrophic growth of Trichoderma species in peat. Using mAbs specific to the hemicellulase enzyme ,-galactosidase (AGL) and the cellulase enzyme endo-1,4-,-glucanase (EG), the relationship between the saprotrophic growth dynamics of a biocontrol strain of Trichoderma hamatum and the concomitant production of these enzymes in peat-based microcosms was studied. Enzyme activity assays and enzyme protein concentrations derived by enzyme-linked immunosorbent assay (ELISA) established the precision and sensitivity of mAb-based assays in quantifying enzyme production during active growth of the fungus. Trends in enzyme activities and protein concentrations were similar for both enzymes, during a 21-day sampling period in which active growth and sporulation of the fungus in peat was quantified using an independent mAb-based assay. There was a sharp increase in active biomass of T. hamatum 3 days after inoculation of microcosms with phialoconidia. After 3 days there was a rapid decline in active biomass which coincided with sporulation of the fungus. A similar trend was witnessed with EG activities and concentrations. This showed that EG production related directly to active growth of the fungus. The trend was not found, however, with AGL. There was a rapid increase in enzyme activities and protein concentrations on day 3, after which they remained static. The reason for the maintenance of elevated AGL probably resulted from secretion of the enzyme from conidia and chlamydospores. ELISA, immunofluoresence and immunogold electron microscopy studies of these cells showed that the enzyme is localized within the cytoplasm and is secreted extracellularly into the surrounding environment. It is postulated that release of oligosaccharides from polymeric hemicellulose by the constitutive spore-bound enzyme leads to AGL induction and could act as an environmental cue for spore germination. [source] Bacterial quorum sensing and nitrogen cycling in rhizosphere soilFEMS MICROBIOLOGY ECOLOGY, Issue 2 2008Kristen M. DeAngelis Abstract Plant photosynthate fuels carbon-limited microbial growth and activity, resulting in increased rhizosphere nitrogen (N) mineralization. Most soil organic nitrogen is macromolecular (chitin, protein, nucleotides); enzymatic depolymerization is likely rate limiting for plant nitrogen accumulation. Analyzing Avena (wild oat) planted in microcosms containing sieved field soil, we observed increased rhizosphere chitinase and protease-specific activities, bacterial cell densities, and dissolved organic nitrogen (DON) compared with bulk soil. Low-molecular-weight (MW) DON (<3000 Da) was undetectable in bulk soil but comprised 15% of rhizosphere DON. Extracellular enzyme production in many bacteria requires quorum sensing (QS), cell-density-dependent group behavior. Because proteobacteria are considered major rhizosphere colonizers, we assayed the proteobacterial QS signals N -acyl-homoserine lactones (AHLs), which were significantly increased in the rhizosphere. To investigate the linkage between soil signaling and nitrogen cycling, we characterized 533 bacterial isolates from Avena rhizosphere: 24% had chitinase or protease activity and AHL production; disruption of QS in seven of eight isolates disrupted enzyme activity. Many Alphaproteobacteria were newly found with QS-controlled extracellular enzyme activity. Enhanced specific activities of nitrogen-cycling enzymes accompanied by bacterial density-dependent behaviors in rhizosphere soil gives rise to the hypothesis that QS could be a control point in the complex process of rhizosphere nitrogen mineralization. [source] Novel domains of the prokaryotic two-component signal transduction systemsFEMS MICROBIOLOGY LETTERS, Issue 1 2001Michael Y. Galperin Abstract The archetypal two-component signal transduction systems include a sensor histidine kinase and a response regulator, which consists of a receiver CheY-like domain and a DNA-binding domain. Sequence analysis of the sensor kinases and response regulators encoded in complete bacterial and archaeal genomes revealed complex domain architectures for many of them and allowed the identification of several novel conserved domains, such as PAS, GAF, HAMP, GGDEF, EAL, and HD-GYP. All of these domains are widely represented in bacteria, including 19 copies of the GGDEF domain and 17 copies of the EAL domain encoded in the Escherichia coli genome. In contrast, these novel signaling domains are much less abundant in bacterial parasites and in archaea, with none at all found in some archaeal species. This skewed phyletic distribution suggests that the newly discovered complexity of signal transduction systems emerged early in the evolution of bacteria, with subsequent massive loss in parasites and some horizontal dissemination among archaea. Only a few proteins containing these domains have been studied experimentally, and their exact biochemical functions remain obscure; they may include transformations of novel signal molecules, such as the recently identified cyclic diguanylate. Recent experimental data provide the first direct evidence of the participation of these domains in signal transduction pathways, including regulation of virulence genes and extracellular enzyme production in the human pathogens Bordetella pertussis and Borrelia burgdorferi and the plant pathogen Xanthomonas campestris. Gene-neighborhood analysis of these new domains suggests their participation in a variety of processes, from mercury and phage resistance to maintenance of virulence plasmids. It appears that the real picture of the complexity of phosphorelay signal transduction in prokaryotes is only beginning to unfold. [source] Reliable high-throughput screening with Pichia pastoris by limiting yeast cell death phenomenaFEMS YEAST RESEARCH, Issue 2 2004Roland Weis Abstract Comparative screening of gene expression libraries employing the potent industrial host Pichia pastoris for improving recombinant eukaryotic enzymes by protein engineering was an unsolved task. We simplified the protocol for protein expression by P. pastoris and scaled it down to 0.5-ml cultures. Optimising standard growth conditions and procedures, programmed cell death and necrosis of P. pastoris in microscale cultures were diminished. Uniform cell growth in 96-deep-well plates now allows for high-throughput protein expression and screening for improved enzyme variants. Furthermore, the change from one host for protein engineering to another host for enzyme production becomes dispensable, and this accelerates the protein breeding cycles and makes predictions for large-scale production more accurate. [source] Seasonal variation in enzyme activities and temperature sensitivities in Arctic tundra soilsGLOBAL CHANGE BIOLOGY, Issue 7 2009MATTHEW D. WALLENSTEIN Abstract Arctic soils contain large amounts of organic matter due to very slow rates of detritus decomposition. The first step in decomposition results from the activity of extracellular enzymes produced by soil microbes. We hypothesized that potential enzyme activities are low relative to the large stocks of organic matter in Arctic tundra soils, and that enzyme activity is low at in situ temperatures. We measured the potential activity of six hydrolytic enzymes at 4 and 20 °C on four sampling dates in tussock, intertussock, shrub organic, and shrub mineral soils at Toolik Lake, Alaska. Potential activities of N -acetyl glucosaminidase, ,-glucosidase, and peptidase tended to be greatest at the end of winter, suggesting that microbes produced enzymes while soils were frozen. In general, enzyme activities did not increase during the Arctic summer, suggesting that enzyme production is N-limited during the period when temperatures would otherwise drive higher enzyme activity in situ. We also detected seasonal variations in the temperature sensitivity (Q10) of soil enzymes. In general, soil enzyme pools were more sensitive to temperature at the end of the winter than during the summer. We modeled potential in situ,-glucosidase activities for tussock and shrub organic soils based on measured enzyme activities, temperature sensitivities, and daily soil temperature data. Modeled in situ enzyme activity in tussock soils increased briefly during the spring, then declined through the summer. In shrub soils, modeled enzyme activities increased through the spring thaw into early August, and then declined through the late summer and into winter. Overall, temperature is the strongest factor driving low in situ enzyme activities in the Arctic. However, enzyme activity was low during the summer, possibly due to N-limitation of enzyme production, which would constrain enzyme activity during the brief period when temperatures would otherwise drive higher rates of decomposition. [source] Rennin-like milk coagulant enzyme produced by a local isolate of MucorINTERNATIONAL JOURNAL OF DAIRY TECHNOLOGY, Issue 4 2003Z A Tubesha Among 20 isolates of Mucor isolated from various environments in Jordan and found to produce a rennin-like acid protease, known as Mucor rennin-like enzyme (MRE), Mucor J20 was found to produce the highest level of MRE. The optimum incubation conditions for enzyme production in a fortified wheat bran mixture using solid-state fermentation were 3,4 days at 30°C. The highest MRE activity (185,200 rennin units or RU) was produced in a medium containing wheat bran and lentil straw (1 : 1 w/w) moistened with whey, and incubated in clay pots at 30°C for 4 days. A slightly lower activity value (178 RU) was found when using a mineral salt solution or distilled water instead of whey, or when using wheat bran alone with whey. At pH 4, the MRE retained its complete activity (100%) for 6 weeks at 5°C and 10°C, and for 3 and 2 weeks at 20°C and 30°C, respectively. After heating at 60°C for 10 min, the enzyme lost its activity at all pH levels used (pH 2,8). The crude extract of MRE was successfully applied in the manufacture of a cheese curd. [source] Phytase production by Sporotrichum thermophile in a cost-effective cane molasses medium in submerged fermentation and its application in breadJOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2008B. Singh Abstract Aims:, Phytase production by Sporotrichum thermophile in a cost-effective cane molasses medium in submerged fermentation and its application in bread. Methods and Results:, The production of phytase by a thermophilic mould S. thermophile was investigated using free and immobilized conidiospores in cane molasses medium in shake flasks, and stirred tank and air-lift fermenters. Among surfactants tested, Tweens (Tween-20, 40 and 80) and sodium oleate increased phytase accumulation, whereas SDS and Triton X-100 inhibited the enzyme production. The mould produced phytase optimally at aw 0·95, and it declined sharply below this aw value. The enzyme production was comparable in air-lift and stirred tank reactors with a marked reduction in fermentation time. Among the matrices tried, Ca-alginate was the best for conidiospore immobilization, and fungus secreted sustained levels of enzyme titres over five cycles. The phytic acid in the dough was efficiently hydrolysed by the enzyme accompanied by the liberation of soluble phosphate in the bread. Conclusions:, The phytase production by S. thermophile was enhanced in the presence of Tween-80 in cane molasses medium. A peak in enzyme production was attained in 48 h in the fermenter when compared with that of 96 h in shake flasks. Ca-alginate immobilized conidiospores germinated to produce fungal growth that secreted sustained levels of phytase over five cycles. The bread made with phytase contained reduced level of phytic acid and a high-soluble phosphate. Significance and Impact of the Study:, The phytase accumulation by S. thermophile was increased by the surfactants. The sustainability of enzyme production in stirred tank and air-lift fermenters suggested the possibility for scaling up of phytase. The bread made with phytase contained low level of antinutrient, i.e. phytic acid. [source] pH Control of the production of recombinant glucose oxidase in Aspergillus nidulansJOURNAL OF APPLIED MICROBIOLOGY, Issue 2 2004R. Luque Abstract Aims:, Recombinant Aspergillus nidulans sVAL040, capable of synthesizing and secreting glucose oxidase derived from Aspergillus niger was used to study the influence of pH and carbon source on enzyme production. Methods and Results:, Glucose oxidase gene (goxC) was expressed under transcriptional regulation by using the promoter of A. nidulans xlnB gene (encoding an acidic xylanase). A maximum specific glucose oxidase activity of approx. 10 U mg,1 protein and a maximum volumetric productivity of 29·9 U l,1 h,1 were obtained at pH 5·5, after 80 h of growth by using xylose as inducer. Enzyme volumetric productivity increased when xylans were used instead of xylose; however, specific glucose oxidase activity did not differ significantly. Conclusions:, Specific GOX activity obtained at pH 5·5 are two to three times more than those previously described for goxC multicopy transformants of A. nidulans. Xylans were a more powerful inducer than xylose although fungal growth was lower when the polymers were used. Significance and Impact of the Study:, The obtained results by using xlnB promoter in A. nidulans could be useful in improving heterologous enzyme production by using genetic- and process-engineering strategies. [source] The influence of extracellular H2O2 production on decolorization ability in fungiJOURNAL OF BASIC MICROBIOLOGY, Issue 6 2006Ivana Eichlerová Dr. A set of 50 randomly chosen fungal strains belonging to different basidiomycete species was tested for H2O2 and ligninolytic enzyme production and for decolorization of synthetic dyes Orange G and Remazol Brilliant Blue R. The decolorization capacity of individual strains was influenced by the level of H2O2 and laccase activity. The strains producing H2O2 at a concentration of 1.0,1.5 µM exhibited the most efficient decolorization; higher or lower H2O2 concentration reduced this ability. None of the strains without a detectable laccase activity was able to decolorize the tested dyes. (© 2006 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] Lactic acid fermentation of food waste using integrated glucoamylase productionJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 1 2009Xiao Qiang Wang Abstract Commercial enzyme is usually needed for the bioconversion of organic waste or biomass. The overall cost could be reduced very significantly if enzyme production could be integrated with its application, avoiding unnecessary steps in enzyme production (such as concentration, recovery and transportation). This investigation attempted to integrate crude glucoamylase production with lactic acid fermentation of food waste. A maximum glucoamylase activity of 1850 U g,1 was obtained with Aspergillus nigerduring solid-state fermentation (SSF) of food waste, 14.8 times more than that obtained during submerged fermentation (SmF). The optimum pH for producing glucoamylase was 4.6, and glucoamylase retained 83.5% of peak activity at pH 3.0. Without any recovery treatment, the glucoamylase produced by SSF could be used directly for lactic acid fermentation of food waste. Lactic acid concentration reached 45.5 g L,1 with the addition of the crude enzyme, 72% higher than the control. No side-effects were caused by the viable A. niger in the crude enzyme. This work successfully integrated glucoamylase production with lactic acid fermentation. The enzyme produced by SSF of food waste had sufficient activity to be used directly without any treatment. The integrated process proposed in this study was very economical and may be helpful to other bioconversions. Copyright © 2008 Society of Chemical Industry [source] Isolation and characterization of strains of Flavobacterium columnare from BrazilJOURNAL OF FISH DISEASES, Issue 4 2005H C P Figueiredo Abstract Flavobacterium columnare is an important pathogen of freshwater fish, implicated in skin and gill disease, often causing high mortality. An outbreak of skin disease in fingerling and adult Nile tilapia, Oreochromis niloticus (L.), cultivated in a recirculation system, was investigated. Four strains were isolated and characterized by biochemical reactions, enzyme production, fatty acid profile and analysis of the 16S-23S rDNA intergenic spacer region. All strains were identified as F. columnare. Experimental infection assays with one of these strains (BZ-5-02) were conducted and pathogenicity (by intramuscular route) was demonstrated in Nile tilapia and channel catfish, Ictalurus punctatus (Rafinesque). This is the first report of characterization of Brazilian strains of F. columnare. [source] ROLE OF SATURATED FATTY ACIDS IN LIPASE PRODUCTION , USING PSEUDOMONAS AERUGINOSAJOURNAL OF FOOD BIOCHEMISTRY, Issue 6 2007A.N. SARAVANAN ABSTRACT Complex substrates always induce substantial amount of enzyme production during hydrolysis by microorganisms. In this study, ghee was taken for its saturated fatty acid content and analyzed as an inducer for the production of lipase. With ghee emulsion, the bacterium Pseudomonas aeruginosa at optimal condition produced 60 units/min/L at 72 h. With olive oil emulsion, this organism produced only 41 units/min/L as maximum at 96 h. The saturated fatty acids present in ghee make it a hard substance for hydrolysis, which is the reason for the increased enzyme production. This was evaluated by the iodine number experiment. Ghee can also reduce the production cost whereas the costlier olive oil constitutes 25,50% of the total production cost for a commercial scale. The experimental results showed that the saturated fatty acids play an important role in lipase enzyme induction by P. aeruginosa. The use of ghee is cost-effective; hence, it can be used as a potential inducer for lipase production. PRACTICAL APPLICATIONS Lipases are industrially very important enzymes. They are used in pharmaceutical, food, soap and other industries. In lipase production, olive oil is the main constituent. Comparatively, olive oil is costlier; hence, it increases the production cost of lipase. So, this study was done to replace olive oil with a much cheaper ghee using Pseudomonas aeruginosa. The ghee-containing medium gave a very good result because of the presence of complex saturated fatty acids. The ghee-containing medium produced 60 units/min/L at 72 h. The olive oil medium, which contains mainly unsaturated fatty acids, produced only 41 units/min/L as maximum at 96 h. Hence, in the commercial scale, ghee can reduce raw material cost as well as operation time cost significantly when it is used as substrate. [source] ISOLATION AND IDENTIFICATION OF A NOVEL ASPERGILLUS JAPONICUS JN19 PRODUCING ,-FRUCTOFURANOSIDASE AND CHARACTERIZATION OF THE ENZYMEJOURNAL OF FOOD BIOCHEMISTRY, Issue 6 2006LI-MEI WANG ABSTRACT A novel strain, Aspergillus sp. JN19, producing,-fructofuranosidase (FFase), was isolated from soil. According to the physiological and biochemical characteristics and its 18S rDNA gene sequence analysis, it was identified as Aspergillus japonicus. The optimal conditions for production of fructofuranosidase by A. japonicus JN-19 were investigated. The initial concentration of sucrose was 15 to 18%. Yeast extract was the best nitrogen source. K2HPO4 was effective in increasing enzyme production. The enzyme activity was increased to about 1.3 times by addition of 0.2% carboxymethylcellulose in the medium. The highest FFase activity was 55.42 U/mL at pH 5.5 and 30C, and production yield of fructooligosaccharides was 55.8%. Some characteristics of purified FFase were also studied. [source] Chitinolytic activity of endophytic Streptomyces and potential for biocontrolLETTERS IN APPLIED MICROBIOLOGY, Issue 6 2008M.C. Quecine Abstract Aims:, Biological sources for the control of plant pathogenic fungi remain an important objective for sustainable agricultural practices. Actinomycetes are used extensively in the pharmaceutical industry and agriculture owing to their great diversity in enzyme production. In the present study, therefore, we evaluated chitinase production by endophytic actinomycetes and the potential of this for control of phytopathogenic fungi. Methods and Results:, Endophytic Streptomyces were grown on minimum medium supplemented with chitin, and chitinase production was quantified. The strains were screened for any activity towards phytopathogenic fungi and oomycetes by a dual-culture in vitro assay. The correlation between chitinase production and pathogen inhibition was calculated and further confirmed on Colletotrichum sublineolum cell walls by scanning electron microscopy. Conclusions:, This paper reports a genetic correlation between chitinase production and the biocontrol potential of endophytic actinomycetes in an antagonistic interaction with different phytopathogens, suggesting that this control could occur inside the host plant. Significance and Impact of the Study:, A genetic correlation between chitinase production and pathogen inhibition was demonstrated. Our results provide an enhanced understanding of endophytic Streptomyces and its potential as a biocontrol agent. The implications and applications of these data for biocontrol are discussed. [source] Culture of Staphylococcus xylosus in fish processing by-product-based media for lipase productionLETTERS IN APPLIED MICROBIOLOGY, Issue 6 2008F. Ben Rebah Abstract Aims:, The objective of this study was to demonstrate that fish-processing by-products could be used as sole raw material to sustain the growth of Staphylococcus xylosus for lipase production. Methods and Results:, Bacterial growth was tested on supernatants generated by boiling (100°C for 20 min) of tuna, sardine, cuttlefish and shrimp by-products from fish processing industries. Among all samples tested, only supernatants generated from shrimp and cuttlefish by-products sustained the growth of S. xylosus. Shrimp-based medium gave the highest growth (A600 = 22) after 22 h of culture and exhibited the maximum lipase activity (28 U ml,1). This effect may be explained by better availability of nutrients, especially, in shrimp by-products. Standard medium (SM) amendments to sardine and tuna by-product-based media stimulated the growth of S. xylosus and the highest A600 values were obtained with 75% SM. Lipase activity, however, remained below 4 U ml,1 for both sardine and tuna by-product-based media. Conclusions:, Fish by-products could be used for the production of highly valuable enzymes. Significance and Impact of the Study:, The use of fish by-products in producing S. xylosus- growth media can reduce environmental problems associated with waste disposal and, simultaneously, lower the cost of biomass and enzyme production. [source] Improved polygalacturonase production from Bacillus sp.LETTERS IN APPLIED MICROBIOLOGY, Issue 5 2002MG-cp-2 under submerged (SmF), solid state (SSF) fermentation Aims:,To investigate the effect of amino acids, vitamins and surfactants on polygalacturonase production from Bacillus sp. MG-cp-2 under submerged (SmF) and solid state fermentation (SSF). Methods and Results:,Bacillus sp. MG-cp-2 was isolated from the outer covering of the seeds of Celastrus paniculatus. Out of the various surfactants, amino acids and vitamins, Tween-60, DL -serine and folic acid maximally enhanced polygalacturonase production by 2·7-fold (240·0 U ml,1), 4·0-fold (360·0 U ml,1) and 3·8-fold (342·0 U ml,1) respectively, under submerged fermentation (SmF). In solid state fermentation (SSF), Tween-80, pyridoxine and DL -ornithine monohydrochloride induced highest enzyme production up to 1·73-fold (6956·5 U g,1), 5·3-fold (21224·4 U g,1) and 5·74-fold (23076·9 U g,1), respectively. Conclusion:,Amino acids and their analogues, vitamins and surfactants effect significantly polygalacturonase production by Bacillus sp. MG-cp-2 when grown under submerged (SmF) and solid state fermentation (SSF) conditions. Significance and Impact of the Study:,The study provides useful information about regulation of polygalacturonase biosynthesis in Bacillus sp. MG-cp-2, which appears to be an interplay of nutritional and physical factors. Alkaline polygalacturonase from Bacillus sp. MG-cp-2 will be extremely useful in the treatment of alkaline pectic waste waters from vegetable and fruit processing industries and in degumming of bast fibres. [source] The secretome of Pleurotus sapidusPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 18 2005Holger Zorn Dr. Abstract Due to their unique capability to attack lignified biopolymers, extracellular enzymes of white-rot fungi enjoy an increasing interest in various fields of white biotechnology. The edible fungus Pleurotus sapidus was selected as a model organism for the analysis of the secretome by means of 2-DE. For enzyme production, the fungus was grown in submerged cultures either on peanut shells or on glass wool as a carrier material. Identification of the secreted enzymes was performed by tryptic digestion, ESI-MS/MS ab initio sequencing, and homology searches against public databases. The spectrum of secreted enzymes comprised various types of hydrolases and lignolytic enzymes of the manganese peroxidase/versatile peroxidase family. While peptidases were secreted mainly by the cultures grown on peanut shells, versatile peroxidase type enzymes dominated in the cultures grown on glass wool. [source] Environmental performance of lignocellulosic bioethanol production from Alfalfa stemsBIOFUELS, BIOPRODUCTS AND BIOREFINING, Issue 2 2010Sara González-García Abstract A ,well-to-wheel' analysis was conducted for bioethanol obtained from alfalfa stems by means of the Life Cycle Assessment (LCA) methodology. This analysis was compared with two blends of conventional gasoline with bioethanol (E10 and E85), all used in a mid-size car. A biochemical process including enzymatic hydrolysis and simultaneous saccharification and fermentation was considered. The life cycles of the fuels include gasoline production, alfalfa agriculture, lignocellulosic bioethanol production, blend production, and finally the use of fuels. The production of the alfalfa plant has two products: high-protein leaves for animal feed (the main driving force) and high-fiber stems. In this study, we assumed two allocation procedures based on mass and protein content, the latter reflecting the greater value of the leaves. According to the results, the use of bioethanol-based fuels leads to reduced global warming potential. A reduction in fossil fuel extraction of up to 72% could be achieved when pure bioethanol is used as transport fuel. On the contrary, bioethanol fuels are not the most suitable option when assessing acidification, eutrophication, and photochemical oxidant formation impact categories, mainly due to the higher impact from the upstream processes (specifically agricultural activities). LCA methodology helped to identify the key areas in the bioethanol production where researchers and technicians need to work to improve the environmental performance, paying special attention to enzyme production, onsite energy generation and distillation processes as well as agricultural activities. Copyright © 2010 Society of Chemical Industry and John Wiley & Sons, Ltd [source] High-level bacterial cellulase accumulation in chloroplast-transformed tobacco mediated by downstream box fusionsBIOTECHNOLOGY & BIOENGINEERING, Issue 4 2009Benjamin N. Gray Abstract The Thermobifida fusca cel6A gene encoding an endoglucanase was fused to three different downstream box (DB) regions to generate cel6A genes with 14 amino acid fusions. The DB-Cel6A fusions were inserted into the tobacco (Nicotiana tabacum cv. Samsun) chloroplast genome for protein expression. Accumulation of Cel6A protein in transformed tobacco leaves varied over approximately two orders of magnitude, dependent on the identity of the DB region fused to the cel6A open reading frame (ORF). Additionally, the DB region fused to the cel6A ORF affected the accumulation of Cel6A protein in aging leaves, with the most effective DB regions allowing for high level accumulation of Cel6A protein in young, mature, and old leaves, while Cel6A protein accumulation decreased with leaf age when less effective DB regions were fused to the cel6A ORF. In the most highly expressed DB-Cel6A construct, enzymatically active Cel6A protein accumulated at up to 10.7% of total soluble leaf protein (%TSP). The strategy used for high-level endoglucanase expression may be useful for expression of other cellulolytic enzymes in chloroplasts, ultimately leading to cost-effective heterologous enzyme production for cellulosic ethanol using transplastomic plants. Biotechnol. Bioeng. 2009;102: 1045,1054. © 2008 Wiley Periodicals, Inc. [source] Expression of Aspergillus hemoglobin domain activities in Aspergillus oryzae grown on solid substrates improves growth rate and enzyme productionBIOTECHNOLOGY JOURNAL, Issue 7-8 2006Rob te Biesebeke Dr. Abstract DNA fragments coding for hemoglobin domains (HBD) were isolated from Aspergillus oryzae and Aspergillus niger. The HBD activities were expressed in A. oryzae by introduction of HBD gene fragments under the control of the promoter of the constitutively expressed gpdA gene. In the transformants, oxygen uptake was significantly higher, and during growth on solid substrates the developed biomass was at least 1.3 times higher than that of the untransformed wild-type strain. Growth rate of the HBD-activity-producing strains was also significantly higher compared to the wild type. During growth on solid cereal substrates, the amylase and protease activities in the extracts of the HBD-activity-producing strains were 30,150% higher and glucoamylase activities were at least 9 times higher compared to the wild-type strain. These results suggest that the Aspergillus HBD-encoding gene can be used in a self-cloning strategy to improve biomass yield and protein production of Aspergillus species. [source] Multicomponent cellulase production by Cellulomonas biazotea NCIM-2550 and its applications for cellulosic biohydrogen productionBIOTECHNOLOGY PROGRESS, Issue 2 2010Ganesh D. Saratale Abstract Among four cellulolytic microorganisms examined, Cellulomonas biazotea NCIM-2550 can grow on various cellulosic substrates and produce reducing sugar. The activity of cellulases (endoglucanase, exoglucanase, and cellobiase), xylanase, amylase, and lignin class of enzymes produced by C. biazotea was mainly present extracellularly and the enzyme production was dependent on cellulosic substrates (carboxymethyl cellulose [CMC], sugarcane bagasse [SCB], and xylan) used for growth. Effects of physicochemical conditions on cellulolytic enzyme production were systematically investigated. Using MnCl2 as a metal additive significantly induces the cellulase enzyme system, resulting in more reducing sugar production. The efficiency of fermentative conversion of the hydrolyzed SCB and xylan into clean H2 energy was examined with seven H2 -producing pure bacterial isolates. Only Clostridiumbutyricum CGS5 exhibited efficient H2 production performance with the hydrolysate of SCB and xylan. The cumulative H2 production and H2 yield from using bagasse hydrolysate (initial reducing sugar concentration = 1.545 g/L) were approximately 72.61 mL/L and 2.13 mmol H2/g reducing sugar (or 1.91 mmol H2/g cellulose), respectively. Using xylan hydrolysate (initial reducing sugar concentration = 0.345 g/L) as substrate could also attain a cumulative H2 production and H2 yield of 87.02 mL/L and 5.03 mmol H2/g reducing sugar (or 4.01 mmol H2/g cellulose), respectively. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010 [source] Optimization of Cyclodextrin Glycosyltransferase Production from KlebsiellapneumoniaeAS-22 in Batch, Fed-Batch, and Continuous CulturesBIOTECHNOLOGY PROGRESS, Issue 6 2003Bharat N. Gawande Production of a novel cyclodextrin glycosyltransferase (CGTase) from Klebsiella pneumoniaeAS-22 strain, which converts starch predominantly to ,-CD at high conversion yields, in batch, fed-batch, and continuous cultures, is presented. In batch fermentations, optimization of different operating parameters such as temperature, pH, agitation speed, and carbon-source concentration resulted in more than 6-fold increase in CGTase activity. The enzyme production was further improved by two fed-batch approaches. First, using glucose-based feed to increase cell density, followed by starch-based feed to induce enzyme production, resulted in high cell density of 76 g dry cell weight/L, although the CGTase production was low. Using the second approach of a single dextrin-based feed, 20-fold higher CGTase was produced compared to that in batch fermentations with media containing tapioca starch. In continuous operation, more than 8-fold increase in volumetric CGTase productivity was obtained using dextrin-based media compared to that in batch culture using starch-based media. [source] Design and Installation of a Next Generation Pilot Scale Fermentation SystemBIOTECHNOLOGY PROGRESS, Issue 3 2003B. Junker Four new fermenters were designed and constructed for use in secondary metabolite cultivations, bioconversions, and enzyme production. A new PC/PLC-based control system also was implemented using GE Fanuc PLCs, Genius I/O blocks, and Fix Dynamics SCADA software. These systems were incorporated into an industrial research fermentation pilot plant, designed and constructed in the early 1980s. Details of the design of these new fermenters and the new control system are described and compared with the existing installation for expected effectiveness. In addition, the reasoning behind selection of some of these features has been included. Key to the design was the goal of preserving similarity between the new and previously existing and successfully utilized fermenter hardware and software installations where feasible but implementing improvements where warranted and beneficial. Examples of enhancements include strategic use of Inconel as a material of construction to reduce corrosion, piping layout design for simplified hazardous energy isolation, on-line calculation and control of nutrient feed rates, and the use of field I/O modules located near the vessel to permit low-cost addition of new instrumentation. [source] Production of a Polyester Degrading Extracellular Hydrolase from Thermomonospora fuscaBIOTECHNOLOGY PROGRESS, Issue 5 2002Mona K. Gouda The production of a polyester-degrading hydrolase from the thermophilic actinomycete Thermomonospora fusca was investigated with regard to its potential technical application. Only in the presence of a polyester (random aliphatic-aromatic copolyester from 1,4-butanediol, terephthalic acid, and adipic acid with around 40,50 mol % terephthalic acid in the acid component), the excretion of the extracellular enzyme could be achieved with an optimized synthetic medium using pectin and NH4Cl as nitrogen source. Compared to complex media, a significantly higher specific activity at comparable volumetric yields could be obtained, thus reducing the expenditure for purification. The activity profile in the medium is controlled by a complex process involving (1) induction of enzyme excretion, (2) enzyme adsorption on the hydrophobic polyester surface, (3) inhibition of enzyme generation by monomers produced by polyester cleavage, and (4) enzyme denaturation. Diafiltration with cellulose acetate membranes as the sole downstream processing step led to a product of high purity and with sufficient yield (60% of total activity). Scaling-up from shaking flasks to a fermentor scale of 100 L revealed no specific problems. However, the excretion of the hydrolase by the actinomycete turned out to be inhibited by the degradation products (monomers) of the aliphatic-aromatic copolyester used as inductor for the enzyme production. The crude enzyme exhibited generally similar properties (temperature and pH optimum) as the highly purified hydrolase described previously; however, the storage capability and thermal stability is improved when the crude enzyme solution is diafiltrated. [source] An Approach for Enhancing Heterologous Production of Providencia Rettgeri Penicillin Acylase in Escherichia coliBIOTECHNOLOGY PROGRESS, Issue 3 2000C. Perry Chou Heterologous production of Providencia rettgeri penicillin acylase (PAC) was optimized inEscherichia coli. Several factors, including carbon, temperature, and host effects, were identified to be critical for the enzyme overproduction. The optimum culture conditions for the enzyme production vary for different host/vector systems. With the optimization, both volumetric and specific PAC activities could be significantly improved by more than 50-fold compared to the native expression in P. rettgeri. The heterologous production could be possibly limited by translation or posttranslational steps, depending on the culture temperature and host/vector system. To our knowledge, this is the first evidence demonstrating the limiting step for the production of P. rettgeri PAC and the existence of the P. rettgeri PAC precursor. [source] Candida albicans proteinases and host/pathogen interactionsCELLULAR MICROBIOLOGY, Issue 10 2004Julian Naglik Summary Candida infections are common, debilitating and often recurring fungal diseases and a problem of significant clinical importance. Candida albicans, the most virulent of the Candida spp., can cause severe mucosal and life-threatening systemic infections in immunocompromised hosts. Attributes that contribute to C. albicans virulence include adhesion, hyphal formation, phenotypic switching and extracellular hydrolytic enzyme production. The extracellular hydrolytic enzymes, especially the secreted aspartyl proteinases (Saps), are one of few gene products that have been shown to directly contribute to C. albicans pathogenicity. Because C. albicans is able to colonize and infect almost every tissue in the human host, it may be crucial for the fungus to possess a number of similar but independently regulated and functionally distinct secreted proteinases to provide sufficient flexibility in order to survive and promote infection at different niche sites. The aim of this review is to explore the functional roles of the C. albicans proteinases and how they may contribute to the host/pathogen interaction in vivo. [source] | ||||||||||||||||||||||||||||||||||