Beta-glucanase Activity Research Articles

Glucose-responsive carboxymethyl chitosan/ sodium alginate film protects against mechanical wound on postharvest blueberry

Fresh fruits and vegetables, particularly berries, are highly susceptible to mechanical damage, which results in nutrient loss, reduced quality, and increased decay from microbial infections, ultimately leading to significant losses. Here, a biodegradable polysaccharide film was employed to encapsulate glucose oxidase (Gox), thereby creating a glucose/wounding-responsive active package that reduced fruit respiration, promoted healing and protected blueberry wounds from mold infection. The combination of carboxymethyl chitosan and sodium alginate resulted in the formation of the polysaccharides film CSF that effectively restricted the permeation of oxygen, water, and carbon dioxide around the surface of the fruit. The encapsulation of Gox in the CSF film (CSFG) reduced glucose levels through enzymatic reaction in surface wounds of the fruit, thereby competing for glucose to establish a starvation-like microenvironment for the pathogenic fungus Botrytis cinerea. The enzymatic reaction also facilitated the production of hydrogen peroxide, which functioned as a fungicide that inhibits the growth of B. cinerea and served as a secondary messenger in fruit immunity. The CSFG film increased beta-glucanase activity, and decreased lipid peroxidation in wounds of blueberries. Therefore, this glucose-responsive CSFG barrier significantly reduced the decay rates of gray mold, caused by B. cinerea in wounded blueberries, by 66.7% compared to the control at 6 d of storage. Taken together, the glucose-responsive active packaging film provides an alternative package for safeguarding blueberries against mechanical damage.

Identification of promising sources of barley (Hordeum vulgare L) for malt beta-glucanase activity and wort beta-glucan content

Five genotypes along with four checks were grown at six locations during the rabi season of 2020-21 and grains were malted and mashed. The genotypes ICARDA-11, ICARDA-9 and DWRB 197 registered the higher malt beta glucanase activity as compared to the checks, with average value more than 370U/kg. Two genotypes, ICARDA-11 and ICARDA-9 were identified with very low wort beta glucans content well below than 200 ppm, the current required levels. These genotypes can be potential sources of these two traits in malt improvement programme of the country.

Isolation, Phenotypic and Molecular Identification of Actinomycetes From Soil and Evaluation of Their Efficiency in Control of the Pathogen Botrytis cinerea Caused Gray Rot Disease on Eggplant

The current study included the isolation of 29 isolates of Actinomycetes from soil samples that collected from different regions of Salah Al-Din Governorate, including districts (Samarra, Tikrit, Balad, Sharqat, Tuz and Ishaqi). The results of the antagonism of these isolates against the pathogenic fungus Botrytis cinerea showed the superiority of isolates S2, S5, Sh3, Sh1 and T2 on other isolates, the inhibition zone of the pathogenic fungus was 2.4, 2.2, 2.2, 2.1 and 1.9 cm, respectively. These isolates were identified using microscopic and biochemical tests, as well as their diagnosis molecularly to the species level using the nucleotide sequence analysis technique of 16S rRNA gene. All isolates belonged to the genus Streptomyces sp., the similarity percentage of the isolates ranged from (99.47-99.62%) with the Streptomyces spp. recorded in the database of the World GenBank. The above isolates were identified as Streptomyces kanamyceticus, S.hygroscopicus, S.achromogenes, S.atratus and S.griseus, respectively, and registered in the database of the World GenBank in NCBI. The filtrates of Streptomyces sp. grown in various media included; Glycerol Tyrosine Medium, Glycerol Yeast Extract Medium, Glycerol Aspargine Medium, Starch-Minerals Medium and Starch Peptone Yeast Extract Medium against pathogenic fungus B. cinerea, the results showed that the S. kanamyceticus filtrate recorded the highest inhibition rate in all media except Starch Mineral medium, which reached 8.13 and 8.66, 4.73, 4.46 and 5.46 mm, respectively. The effectiveness of some active substances with antifungal activity produced from the Streptomyces spp. was estimated, which included the antibiotic Fungichromin, chitinase and beta-glucanase, the results showed the production of fangichromin from S. kanamyceticus, S. hygroscopicus, and S.achromogenes only, which reched to 6.76, 6.66 and 5.4 μg/ml, respectively, on the othe hand, all isolates recorded the activity of chitinase and beta-glucanase, the highest activities were 0.895 and 1.254 units/ml in S. kanamyceticus, respectively.

Genetic variation among selected pure lines from Turkish barley landrace 'Tokak' in yield-related and malting quality traits

Aim of study: Improvement of barley cultivars for malting traits suffers from narrow genetic pool in barley for these traits. Landraces are resources that could be used for this purpose. The present study was conducted to determine the variation for malting quality traits within a Turkish barley landrace. Area of study: The study was undertaken in Tokat, a province in Black Sea Region of Turkey. Material and methods: Twenty-five diverse lines, out of 42 unique genotypes previously identified in ‘Tokak’ landrace (PI 470281) based on DNA markers, were evaluated for malting quality traits along with the malting barley cv. ‘Tokak 157/37’ in four field trials. Thousand-seed weight, test weight, grain yield, lodging, malt extract percentage, diastatic power, alpha amylase and malt beta glucanase activities, malt protein and starch contents were determined. Main results: Principal component analysis of malting quality traits revealed that thousand-seed weight, alpha amylase activity, beta glucanase activity and diastatic power were the most discriminatory traits for the lines. As the average of four trials, 15 of the 25 lines evaluated had higher grain yields and 10 of 25 lines had higher malt extract percentages than the standard cultivar ‘Tokak 157/37’. Malt extract was highest in Line 59 in all environments, and this line also had the highest values for beta glucanase activity and starch content. Line 215 had highest values for alpha amylase activity. Lines 59 and 215 clearly had superior malting quality. Research highlights: These lines could harbor novel alleles for these traits to be used in malting barley improvement.

Constructing and Screening Beta-Glucanase Activity of Metagenomic cDNA Expression Library of Digestive Gland of Achatina fulica

Constructing and Screening Beta-Glucanase Activity of Metagenomic cDNA Expression Library of Digestive Gland of Achatina fulica

Comparative Analysis of the Composition and Properties of Fodder Enzyme Preparations

The composition and properties of a wide variety of domestic and foreign enzyme preparations (EP) used as additives to feeds of farm animals and poultry are analyzed. Content of the main active enzymes – endoglucanases (beta-glucanases), cellobiohydrolases, and xylanases performing biocatalytic destruction of non-starch polysaccharides, which are anti-nutritional factors in feeds causing their incomplete digestion, is determined. It is shown that based on the component composition data and the level of different types of activity the studied enzyme preparations can be classified into three groups: a) with high xylanase and low cellulase (endoglucanase and cellobiohydrolase) content, b) with high cellulase and low xylanase content, c) containing cellobiohydrolases, endoglucanases and xylanases at a different ratio, but without significant prevalence of any of these enzymes. The ability of EP to reduce the viscosity of water-soluble non-starch polysaccharides – xylans and beta-glucans – has been studied. Among the enzyme preparations that have xylanase in their composition and belong to the groups b) and c) a number of preparations have been identified (Econase XT 25, Agroxyl Plus, Agroxyl Premium, Rovabio Max AP, Sunzyme), which reduced the viscosity of aqueous extract of rye containing xylans most effectively when used at the same dosage with regards to xylanase activity. It was shown that xylanase from precisely these EP was not inhibited by the protein inhibitors of rye. The viscosity of water-soluble beta-glucans of barley was most effectively reduced by the EP Xybeten CELL, Cellulase, Agroxyl, Agrocel, Axtra XB 201, Rovabio Max AP, and Vilzim used at the same dosage with regards to beta-glucanase activity. For all the studied EP, no inhibitory effect of the barley extract on beta-glucanase activity was found.

Comparison Analysis of the Composition and Properties of Fodder Enzyme Preparations

The composition and properties of a wide range of domestic and foreign enzyme preparations (EP), used as additives to feeds of farm animals and poultry, are analyzed. The content of the main active enzymes – endoglucanases (beta-glucanases), cellobiohydrolases and xylanases, leading to biocatalytic destruction of non-starch polysaccharides, which are anti-nutritional factors of feeds and causing their incomplete digestion, is determined. It is shown that, based on the data on the component composition and the level of different types of activity, the studied enzyme preparations can be divided into three groups: a) with high xylanase and low cellulase (endoglucanase and cellobiohydrolase) content, b) high cellulase and low xylanase content, c) containing cellobiohydrolases, endoglucanases and xylanases in a different ratio, but without significant predominant of any of these enzymes. The ability of EP to reduce the viscosity of water-soluble non-starch polysaccharides – xylans and beta-glucans- has been studied. Among the enzyme preparations that have xylanase in their composition and belong to groups b) and c), a number of preparations have been determined which, at the same dosage according to xylanase activity, most effectively reduced the viscosity of the aqueous extract of rye containing xylans (Econase XT 25, Agroxyl Plus, Agroxyl Premium, Rovabio Max AP, Sunzyme). It was shown that the xylanase of precisely these EP is not inhibited by protein inhibitors of rye. At the same dosage for beta-glucanase activity, the viscosity of water-soluble beta-glucans of barley was most effectively reduced by the EP Xybeten CELL, Cellulase, Agroxyl, Agrocell, Axtra XB 201, Rovabio Max AP and Vilzyme. For all studied EP, no inhibitory effect of the barley extract on beta-glucanase activity was found.

The Influence of Hot Water and Calcium Chloride on the Changes in Cell Wall Composition and the Activities of Cell Wall Hydrolases during Storage inAgaricus bisporus

Mushrooms show significant losses in firmness due mainly to the changes in cell walls after harvest. This study was performed to determine the effects of CaCl2 and hot water dips on the changes in firmness, cell wall composition and hydrolase activities of mushrooms during storage. Mushrooms were treated with various concentrations of CaCl2 and hot water at various degrees and stored for 12 days at 10C. The results showed that the firmness of mushrooms decreased significantly and their cell walls have undergone significant changes during 12 days storage period after harvest. Polysaccharide content decreased significantly during storage. This decrease was mainly due to the decrease in the strong alkaline-soluble polysaccharides. Xyloglucanase and beta glucanase activities increased significantly during storage, possibly contributing significantly to the polysaccharide losses. CaCl2 and hot water treatments significantly reduced the changes in cell wall components and cell wall hydrolase activities. The results suggest that CaCl2 and hot water treatments could be used to reduce the changes in firmness and cell walls after harvest and extend the shelf life of mushroom. Practical Applications Mushrooms show rapid deterioration after harvest due mainly to changes in cell walls and have a very short shelf life. This significantly reduces their marketing period. The results demonstrate that both hot water and CaCl2 significantly delay cell wall degradation and prolong the shelf life of mushrooms. The results of the study have the potential to be used to decide on what treatments should be used to extend the shelf life and marketing period of mushrooms.

Biocontrol activity of salt tolerant Streptomyces isolates against phytopathogens causing root rot of sugar beet

Biological control of fungi causing root rot on sugar beet by native Streptomyces isolates (C and S2) was evaluated in this study. The dry weight and colony forming unit (CFU) of S2 and C increased when 300 mM NaCl was added to medium. The in vitro antagonism assays showed that both isolates had inhibitory effect against Rhizoctonia solani AG-2, Fusarium solani and Phytophthora drechsleri. In dual culture, Streptomyces isolate C inhibited mycelial growth of R. solani, F. solani and P. drechsleri 45%, 53% and 26%, respectively. NaCl treatment of medium increased biocontrol activity of soluble and volatile compounds of isolate C and S2. After salt treatment, growth inhibition of R. solani, F. solani and P. drechsleri by isolate C increased up to 59%, 70% and 79%, respectively. To elucidate the mode of antagonism, protease, chitinase, beta glucanase, cellulase, lipase and α-amylase activity and siderophore and salicylic acid (SA) production were evaluated. Both isolates showed protease, chitinase and α-amylase activity. Also, biosynthesis of siderophore was detectable for both isolates. Production of siderophore and activity of protease and α-amylase increased after adding salt for both isolates. In contrast, chitinase activity decreased significantly. Production of SA, beta glucanase and lipase by isolate S2 and biosynthesis of cellulase by isolate C were observed in presence and absence of NaCl. Soil treatment with Streptomyces isolate C inhibited root rot of sugar beet caused by P. drechsleri, R. solani and F. solani. Results of this study showed that these two Streptomyces isolates had potential to be utilized as biocontrol agent against fungal diseases especially in saline soils.

A novel chitin‐binding protein fromMoringa oleiferaseed with potential for plant disease control

A thermostable chitin-binding protein (14.3 kDa) with antifungal activity was isolated from Moringa oleifera seeds by affinity chromatography on chitin followed by ion exchange chromatography. NH(2-) CPAIQRCCQQLRNIQPPCRCCQ (Mo-CBP3) is a glycoprotein with 2.5% sugar, pI 10.8, without hemagglutination, chitinase or beta-glucanase activities. Mo-CBP3 possesses in vitro antifungal activity against the phytopathogenicfungi Fusarium solani, F. oxysporum, Colletotrichum musae and C. gloesporioides. Contrarily, Mo-CBP3 did not affect Pythium oligandrum, an oomycete. At 0.05 mg/ml, Mo-CBP3 showed to be fungistatic against F. solani, but at 0.1 mg/ml Mo-CBP3 behaved as a potent fungicidal agent as it inhibited both the spore germination and mycelial growth of F. solani. Surprisingly, the effect of Mo-CBP3 against spore germination was observed even when the protein was heated at 100 degrees C for 1 h or pretreated with 0.15M N-acetyl-D-glucosamine. Mo-CBP3 inhibited the glucose-stimulated acidification of the incubation medium by F. solani. This is apparently caused by structural plasma membrane disarrangement induced by Mo-CBP3. Altogether, these results suggest that Mo-CBP3 might be involved in plant defense mechanisms and could be used as potential antifungal agent for controlling fungal pathogens in plants.

Influence of Beta-glucanase Supplementation on Growth Performances and Digestive Organs Weights of Broiler Chickens Fed Corn, Wheat and Barley-based Diet

3 Abstract: This study was performed with broiler chickens to evaluate the effects of adding beta-glucanase on bird performances and digestive organs weights. A total of 1600 1-day-old broilers (Arbor Acres) were divided into 2 groups and fed either a control diet (group C) based on corn, wheat and barley or a control diet supplemented with beta-glucanase preparation (Safizym GP 40, Lesaffre Feed Additives, France) at the dose of 40 mg per kg of diet (group G). Body weight, feed intake were recorded at 40 days of age. The same day, 8 chickens from each group were slaughtered for carcass yield and digestive organs weights. Experimental data were statistically analyzed by using the ANOVA procedure of Stat View programme. Throughout the whole trial period (1-40 days of age), body weight was significantly (p<0.05) increased (+2.9%) in birds of group G (1723 g) compared to those of group C (1674 g). Moreover, there was no significant difference in feed intake between broilers consuming the control diet supplemented or not with beta-glucanase activity (4052 g). However, the feed conversion ratio was significantly reduced by 4% when the basal diet was supplemented with beta-glucanase (p<0.05). The addition of beta-glucanas e reduced (p<0.05) the small intestine weight related to the body weight compared to C group (5.40 vs.6.98%, respectively). Thus, it is possible to introduce in the broiler diet up to 35% of barley when beta-glucanase is used at th e recommended dose of 40 mg per kg of diet.

β-glucanase Productivity Improvement via Cell Immobilization of Recombinant Escherichia coli Cells in Different Matrices

The studies have been performed to analyze the production of beta-glucanase by a recombinant strain of Escherichia coli immobilized in different matrices. Porous sintered glass SIRAN, Ceramic supporting matrices and Broken Pumice stone as well as SIRAN Raschig-rings were examined for the immobilization of whole bacterial cells. The beta-glucanase activity of bacteria immobilized in CeramTec PST 5 (4-5 mm) was very low. CeramTec PST 5 (1.5-2.5 mm) was found to be the best carrier compared to all other matrices regarding glucanase production (630 U/ml) and compared to enzyme activity produced by free cells (500 U/ml). Different doses of matrices were applied (2, 5, 7, 10 g/lask) in the form of "matrix weight". Using 2 g/flask of CeramTec PST 5 (1.5-2.5 mm) yielded enzyme activity of 630 U/ml). CeramTec gives highest operational stability of beta-glucanase by repeated batch fermentation to 5 cycles, and activity reached 660 U/ml. Scanning electron microscopy observations showed a high number of vegetative cells that continued growth inside the matrices, indicating that beta-glucanase activity improvement was due to the immobilization of the cells.

Effects of nonstarch polysaccharide-hydrolyzing enzymes on performance and amino acid digestibility in turkeys

The effects of a nonstarch polysaccharide-hydrolyzing enzyme product on growth, feed conversion, ME, and digestibility of crude nutrients and amino acids were investigated in a 22-wk trial using male turkeys. Diets were mainly based on wheat, barley, and rye, and the experiment comprised 6 phases. The enzyme product contained endoxylanase and beta-glucanase activity. Fifteen replicated pens were used for studying growth and feed conversion. In 5 pens per treatment, excreta were collected on 5 consecutive days during phases 3 to 6. Ileal digesta was sampled at the end of these phases from 1 bird of each pen. Titanium dioxide was used as indigestible marker to calculate digestibility and ME concentration. In phase 6, but not in the other phases, BW gain and G:F ratio were significantly improved by the supplemented enzyme product. This was associated with a significantly higher ME concentration. With the exception of cystine, no significant effects were measured for the prececal digestibility of amino acids. Amino acid digestibility, calculated based on total excreta, was significantly higher with enzyme supplementation for 12 or 8 out of the 14 analyzed amino acids in phases 5 and 6. In phases 3, 4, and 5, the level of amino acid digestibility was higher by an average of 5 percentage units when calculated on total excreta than on ileal digesta basis. The greatest differences were found for cystine and methionine. Improved digestibilities and energy metabolizability contribute to the effects of nonstarch polysaccharide-hydrolyzing enzymes on feed conversion and growth in heavy turkeys.

Anaerobic digestion of distillery spent wash: Influence of enzymatic pre-treatment of intact yeast cells

The potential benefits of enzymatic digestion of intact yeast cells on anaerobic digestion of Scotch whisky distillery spent wash and pot ale were investigated. Various yeast cell wall hydrolytic enzymes were studied based on their effect on dissolution of cell wall glucan and mannoprotein. The synergistic activity of beta-glucanase and protease showed greater than 90% yeast cell digestion at 37 °C in 24 h. The widely-used industrial enzyme papain showed 95% yeast cell digestion in spent wash at 1% enzyme concentration within 22 h at 50 °C. Anaerobic digestion of pot ale residues containing intact yeast cells pre-treated with lytic enzymes showed COD reductions of 87%, compared with only 13% without enzymes. Similar results were observed with distillery spent wash centrate. The hydrolysis of intact yeast cells in distillery liquid residues was found to be a rate-limiting step in anaerobic treatment of such residues.

EST-SSR markers derived from an elite barley cultivar (Hordeum vulgareL. ‘Morex’): polymorphism and genetic marker potential

Microsatellites or simple sequence repeats have become the markers of choice for marker-assisted selection because of their low template DNA requirement, high reproducibility, and high level of polymorphism. This study investigated a new set of barley (Hordeum vulgare L.) EST-derived SSR markers designed to target gene sequences expressed during grain development, as they are more likely to be important in determining grain quality. The EST sequences (HVSMEh and HVSMEi) were derived from cDNA libraries of the elite six-rowed cultivar Morex, made from spikes harvested at 5 to 45 days after pollination. Approximately half of the 110 SSR markers derived from the ESTs were polymorphic in a panel of 8 diverse barley genotypes, with PIC values between 0.19 and 0.79. Twenty of the new markers were mapped to chromosomal locations using 2 doubled haploid populations. To demonstrate marker potential, quantitative trait locus (QTL) analyses were carried out with phenotypic data on wort beta-glucan content and beta-glucanase activity, two traits with a long history of genetic studies. Most of the EST-SSR markers mapped to within 10 cM of the cellulose synthase (HvCesA) and cellulose synthase-like (HvCslF) genes, which provides highly informative functional markers for tracking these genes in breeding programs. It was also observed that on any given chromosome, the QTL for beta-glucan content and beta-glucanase activity were rarely coincident but tended to occur in adjacent intervals along chromosomal regions, which agreed with their independent genetic basis; the adjacent localization may be important for coordination of cell wall degradation during germination and malting.

Genotypic and environmental variation in grain protein components and their relations to beta-amylase and beta-glucanase activity in malting barley

Eight malting barley cultivars were used to investigate the cultivar and environmental effects on grain protein components and the relationships between protein fractions and β -amylase or β -glucanase activity. The results showed there was a great variation for three protein fraction (albumin, hordein and glutelin) contents over cultivars and locations, and a distinct difference in each protein fraction content between the locations for a given cultivars. Correlation analysis indicated that β -amylase activity was significantly correlated with three protein fraction contents and there was a negative correlation between glutelin content and β -amylase activity, but β -amylase activity positively correlated with albumin or hordein content. Furthermore, there was a significant positive correlation between total protein content and β -glucanase activity, and we found the hordein and glutelin content did not show correlated with β -glucanase activity but the albumin content was a significantly negative correl...

Characterization of a Thermostable and Acidic‐Tolerable β‐Glucanase from Aerobic Fungi Trichoderma koningii ZJU‐T

An extreme thermostable and acidic tolerable beta-glucanase was isolated and characterized from aerobic fungi Trichoderma koningii ZJU-T. The optimal reaction temperature and pH for the beta-glucanase were 100 degrees C and pH 2.0, respectively. The beta-glucanase showed increased stability at higher temperatures and lower pH values when compared to other beta-glucanases. The optimum conditions for the beta-glucanase stability were found to be pH 4.0 and 80 degrees C. Even subjected to 100 degrees C for 3 h, beta-glucanase activity did not show significant reduction. Moreover, K(+) significantly enhanced beta-glucanase activity at the concentration of 1 mM, while EDTA and other metal ions such as Mg(2+), Mn(2+), Zn(2+), Ca(2+), Fe(2+), Pb(2+), and Fe(3+) inhibited beta-glucanase activity. Denaturants, including sodium dodecyl sulfate (SDS) and mercaptoethanol, also inhibited beta-glucanase activity at a concentration of 5%. However, in the presence of 7 M urea, residual activity of the beta-glucanase still remained 14.5%.

Screening and evaluation of the glucoside hydrolase activity in Saccharomyces and Brettanomyces brewing yeasts

The aim of this study was to select and examine Saccharomyces and Brettanomyces brewing yeasts for hydrolase activity towards glycosidically bound volatile compounds. A screening for glucoside hydrolase activity of 58 brewing yeasts belonging to the genera Saccharomyces and Brettanomyces was performed. The studied Saccharomyces brewing yeasts did not show 1,4-beta-glucosidase activity, but a strain dependent beta-glucanase activity was observed. Some Brettanomyces species did show 1,4-beta-glucosidase activity. The highest constitutive activity was found in Brettanomyces custersii. For the most interesting strains the substrate specificity was studied and their activity was evaluated in fermentation experiments with added hop glycosides. Fermentations with Br. custersii led to the highest release of aglycones. Pronounced exo-beta-glucanase activity in Saccharomyces brewing yeasts leads to a higher release of certain aglycones. Certain Brettanomyces brewing yeasts, however, are more interesting for hydrolysis of glycosidically bound volatiles of hops. The release of flavour active compounds from hop glycosides opens perspectives for the bioflavouring and product diversification of beverages like beer. The release can be enhanced by using Saccharomyces strains with high exo-beta-glucanase activity. Higher activities can be found in Brettanomyces species with beta-glucosidase activity.

Protein Haze Formation in White Wines: Effect ofSaccharomyces cerevisiaeCell Wall Components Prepared with Different Procedures

Cell wall material was extracted by five different methods from an oenological strain of Saccharomyces cerevisiae. Enzyme preparations containing beta-glucanase activity (Zymolyase, Glucanex, and Finizym 250L) allowed a better extraction yield compared to that of dithiothreitol (DTT) and EDTA. The yeast extracts were only soluble in part in wine. The wine-soluble fraction (WSF) of the five extracts, differing in both protein and sugar contents, when added in increasing amounts to white wine differently affected protein haze formation, as determined by the heat test, giving dose/response curves of different shapes. The curves obtained with the WSF derived from DDT and Zymolyase extracts showed a plateau value corresponding to 90% and 80% of wine haze reduction, respectively. In contrast, addition of the WSF derived from the other extracts resulted in increased turbidity with respect to the original wine. The mannoproteins (MP), isolated from each yeast extract by Concanavalin-A chromatography, gave dose/response curves showing shapes more similar among them than those obtained with the whole WSFs. The best wine stabilization was obtained with the MP of the DTT and Zymolyase extracts. The supernatants obtained after heating the MP of the different extracts were also tested. The stabilizing effect of the heat-stable MP (HSMP) was always larger than that of the corresponding total (un-heated) MP. The HSMP obtained starting from the DTT and Zymolyase extracts showed the best haze-protecting effect, which was, however, lower than that obtained with their corresponding WSF. This result suggests that wine protein stabilization by compounds of the yeast cell wall could be related, in addition to the action of the MP, also to the presence of other substances of different nature.

Increasing the secretion ability of the kil gene for recombinant proteins in Escherichia coli by using a strong stationary-phase promoter

By using a beta-glucanase from Bacillus as a model protein, we investigated whether the secretion competence based on the action of the kil gene can be improved using stronger promoters for the expression of the kil gene. Since the production of extracellular target proteins also depends on the promoter strengths of the target gene, we constructed four expression vectors with all possible combinations of a weak and a strong stationary-phase promoter for the kil gene, and a weak and a strong constitutive promoter, respectively, for the beta-glucanase gene. The results of batch fermentations showed that the use of stronger promoters generally decreased the cell density. However, a drastic increase of productivity of the cells to produce and secrete beta-glucanase resulted in a significantly higher activity of extracellular beta-glucanase. The yield of extracellular beta-glucanase can be increased (to 168 %) by using a strong promoter for the beta-glucanase alone. However, the increase was much higher when the weak promoter of the kil gene was replaced by a strong stationary-phase promoter (to 221 %). An even higher yield of extracellular beta-glucanase was reached when beta-glucanase was expressed by a strong promoter in addition indicating a combinatorial effect. This shows that the extracellular production of a recombinant target gene can be optimized by tuning the promoter strengths of components, the kil gene and the target gene.