Effect of various sources of nitrogen and carbon on the biosynthesis of proteolytic enzymes in a culture of Aspergillus awamori 21/96

Effects of various conditions of nitrogen and carbon nutrition on the biosynthesis of proteolytic enzymes in a selected culture of Aspergillus awamori 21/96 were studied. This strain was shown to produce proteolytic enzymes constitutively. In the presence of mineral sources of nitrogen, the synthesis of the enzymes under study was not induced by proteinaceous substrates. Optimum conditions of the enzyme biosynthesis were achieved with casein as a source of nitrogen and starch or dulcitol as a source of carbon (which increased the production of the enzymes 1.7 and 8 times, respectively). When the cells were grown on starch, their specific activity exceeded control levels 18 times.     

Characteristics of hydrolase biosynthesis in Bacillus mesentericus grown on various media

The dynamics of the consumption of major carbon and nitrogen sources and the biosynthesis of hydrolytic enzymes were studied in Bacillus mesentericus grown on semisynthetic media. Conditions were chosen that provide the obtaining of the culture liquid with predominantly proteolytic or amylolytic activity. The replacement of maltose with native starch resulted in more intensive accumulation of the biomass and hydrolytic enzymes, and in more rapid (by 3-5 hr) transformation from the logarithmic to the stationary growth phase.     

Enzymes of Candida albicans cell-wall lytic system produced by Streptomyces thermodiastaticus

The production of the enzymes of Candida albicans cell-wall lytic system by S. thermodiastaticus was found to be affected by some growth conditions and nutritional factors. The highest lytic activity was obtained after 18 h of incubation at pH 5.5 and an incubation temperature of 50 degrees C. The carbon source influenced the production of the enzymes of the yeast cell wall lytic system. Maximum lytic activity was obtained when Candida albicans cell-wall (1 g/100 ml) was used as the sole carbon source. NaNO3 at 0.1 g/100 ml level was the best nitrogen source for the biosynthesis of the enzymes of the yeast lytic system. From all phosphor sources, microelements, and growth factors tested, KH2PO4 (1 g/l), ZnSO4 (1 mg/l) and Tween 80 (0.1%), respectively were found to favour highest enzymes production of the lytic system. The Candida albicans cell-wall lytic system produced by S. thermodiastaticus mainly contained chitinolytic and proteolytic activities.     

Influence of different sources of carbon and nitrogen on the biosynthesis of proteolytic complexes by Bacillus circulans 693, Bacillus sp. 27 and Yarrowia lipolytica 2061 strains

The influence of different factors on the biosynthesis of extracellular proteolytic complexes by strains-producers Bacillus circulans 693, Bacillus sp. 27 and Yarrowia lipolytica 2061 at submerged cultivation has been investigated. It has been shown that ammonium hydrocarbonate and gelatin with glucose were optimum carbon and nitrogen sources for synthesis of proteolytic activity of B. circulans strain 693, gelatin with arabinose--for Bacillus sp. 27, gelatin and glycine with sorbose--for Y. lipolytica 2061. It has been established that the cultivation of producers on optimal sources of carbon and nitrogen increased protease activity of cultural liquid of B. circulans 693 3.8 times, Bacillus sp. 27--2.7 times, Y. lipolytica 2061--3.4 times. It has been found that the usage of different protein substrates in cultural medium permitted to obtain the proteolytic enzymes with various specificity with respect to globular and fibrillar proteins.     

绿僵菌产海藻糖水解酶培养条件研究

丝状真菌绿僵菌能产生一系列二糖水解酶,其中包括海藻糖水解酶。这些酶在绿僵菌对昆虫的致病过程中起着重要的作用。本文研究了不同碳源、氮源对金龟子绿僵菌Metarhizium anisopliae var. acridum菌株CQMa102产生与分解昆虫血淋巴中海藻糖等二糖相关的海藻糖水解酶活性的影响。结果表明:分别以葡萄糖、麦芽糖、蔗糖、山梨醇和可溶性淀粉为碳源,金龟子绿僵菌均可产生海藻糖水解酶,但最佳碳源是可溶性淀粉,因为由其诱导产生的海藻糖水解酶具有最高的总活性和比活性以及更多的同工酶,山梨醇次之。硝态氮(NaNO₃)作为唯一氮源时,几乎检测不出海藻糖水解酶活性,而铵态氮((NH₄)₂SO₄)或NaNO₃和有机氮(蛋白胨和酵母浸膏)混合氮源作氮源时,海藻糖水解酶活性都很高。在绿僵菌菌丝提取液和滤液的海藻糖水解酶活性比较中发现:CQMa102在多数碳源的培养基中产生的海藻糖水解酶主要分泌到培养基中,仅有少数结合在细胞壁上。     

Control of the synthesis of plasmin-like and plasminogen-activating proteinases in marine bacteria

The biosynthesis of proteinases with various substrate specificities was studied inBacillus firmus 44b andBacillus oligonitrophilus 21p as influenced by the growth conditions and growth phases of the bacteria. The period of the maximum synthesis of plasmin-like enzymes was observed 6 h later than the period of the maximum growth rate ofB. firmus 44b, and 3 h later than the maximum growth rate period ofB. oligonitrophilus 21 p. The periods of the maximum accumulation of activating enzymes were delayed 9 and 12 h, respectively, as compared to the periods of rapid growth of these two bacteria. Catabolite repression of proteinase synthesis and stimulation of the latter with substrate proteins were insignificant. The production of both plasmin-like and plasminogen-activating enzymes was most sensitive to repression by nitrogen deficiency. The production of plasminogen-activating proteinases was less dependent on the carbon source than the production of plasmin-like enzymes.     

Changes in Starch Formation and Activities of Sucrose Phosphate Synthase and Cytoplasmic Fructose-1,6-bisphosphatase in Response to Source-Sink Alterations

Short term experiments were conducted with vegetative soybean plants (Glycine max L. Merr. `Ransom' or `Arksoy') to determine whether sourcesink manipulations, which rapidly changed the `demand' for sucrose and partitioning of photosynthetically fixed carbon into starch, were associated with alterations in activities of sucrose-P synthase and/or cytoplasmic fructose-1,6-bisphosphatase in leaf extracts. When demand for sucrose from a particular source leaf was increased by defoliation of other source leaves, starch accumulation was restricted and activities of both enzymes were markedly enhanced. When demand for sucrose from source leaves was limited by excision, starch accumulation in the detached leaves was increased while activity of sucrose-P synthase declined sharply. The consistent responsiveness of sucrose-P synthase activity to changes in demand for sucrose supports the contention that regulation of sucrose-P synthase is an integral component of the system which controls sucrose biosynthesis and partitioning of carbon between starch and sucrose biosynthesis in the light.     

Beta-glucanase and chitinase biosynthesis in a culture of a mycophilic strain of Trichoderma viride

The production of extracellular 1,3-, 1,6-beta-glucanases and chitinase was studied during submerged cultivation of a Trichoderma viride strain 3/78 on various carbon sources: glycerol, glucose, lactose, sucrose, laminaran, starch, pustulan, chitin, and Agaricus bisporus fruit bodies. The synthesis of these enzymes and cellulase was studied also under the conditions of depression at low concentrations (10(-2) and 10(-3)M) of the first five aforementioned carbon sources as well as cellobiose, gentiobiose, N-acetyl-beta-D-glucosamine and 0.1% chitooligosaccharides and A. bisporus cell walls. The experiments were conducted with the washed mycelium of this strain grown for 2 days in a medium with glycerol as a carbon source. The results indicated that 1,3- and 1,6-beta-glucanases of the strain were of the constitutive nature and were repressed by such carbon sources as glycerol and glucose. Chitinase and cellulase were shown to be inducible enzymes. Chitinase was induced by N-acetyl-beta-D-glucosamine, chitooligosaccharides and A. bisporus cell walls as well as by lactose when the fungus was grown on this carbon source. Cellulase biosynthesis was induced by lactose, cellobiose and gentiobiose.     

Optimization of culture medium and growth conditions for production of L-arabinose isomerase and D-xylose isomerase by <Emphasis Type="Italic">Lactobacillus bifermentans</Emphasis>

Lactobacillus bifermentans was used to produce the intracellular enzymes L-arabinose isomerase and D-xylose isomerase. Various factors of cultivation (temperature, pH, and incubation period) and culture medium composition (mineral salts, carbon source, and nitrogen source) were studied to select the conditions that maximize production of these enzymes. Arabinose isomerase and xylose isomerase activities were 9.4 and 7.24 U/ml, respectively. They were highest at 9 h of cultivation in the optimized medium, 1.6 times higher than that in the basic MRS broth. The optimal medium composition and cultivation conditions were determined. For optimal growth, the strain required Tween 80 (1 g/l) and a source of inorganic nitrogen (e.g., ammonium citrate). The bacterium had no requirement for sodium acetate for either growth or production of isomerases. The production rate of enzymes was increased when metal ions were added, primarily manganese (2.5 mM). The text was submitted by the authors in English.     

Optimization of growth and hydrolytic enzymes production by Fusarium culmorum using response surface method

Production of hydrolytic enzymes by a phytopathogenic fungus Fusarium culmorum was investigated. The proteolytic activity was observed when the fungus was grown in the medium containing starch or soybean meal as a carbon source. The amylolytic and lipolytic activities were not found. Response surface modeling was applied to shake-flask culture of the fungus to determine the optimum concentration of carbon source and optimal culture time for growth and protease production. The results indicated that the maximum yield of protease production corresponded to the concentration of soybean meal of 1.4?g/ml and culture time of 4.5?days. The fungus growth depends on the concentration of carbon source in the medium whereas the enzyme production was also influenced by the culture time and interaction between these two variables.     

Submerged Cultivation of a Fungal Mutant Strain Humicola Lutea 120-5 Producing Acid Proteinases

The optimal parameters for the cultivation in 10-l fermenters of a mutant strain Humicola lutea 120-5 were established:temperature 30°C, inoculum size 6%, inoculum age 24 h, aeration rate 0,6 vol/vol · min, medium agitation 620 rpm and cultivation time 72 h. A maximal proteolytic activity of 2000 µg tyrosine liberated from 2%casein ml^?1 culture filtrate min^?1 at pH 3.0 and 40°C was obtained under the fixed conditions. α-Amylase biosynthesis during the cultivation of H. lutea 120-5 was observed but it was insignificant to the 72nd h. It is demonstrated that starch can be used as alternative to glucose carbon source. It is proved that the mutant strain H.lutea 120-5 produced two acid proteinases.     

Production of methylanthranilate by the basidiomycete Pycnoporus cinnabarinus (Karst.)

Summary Pyncnoporus cinnabarinus (Polyporaceae) is able to produce methylanthranilate in liquid cultures. Study of the culture conditions of P. cinnabarinus IP I-937 has permitted increase in the aroma productivity by a factor of 16. A low nitrogen concentration, with maltose as carbon source, was required; the culture pH was uncontrolled. The inoculum nature and concentration greatly influence on production: best results were obtained with conidia from a late harvest, used at a rate of 2 × 10⁵ spores/ml. Under these conditions, 18.7 mg methylanthranilate/l was produced after 5 days of culture. Aroma production is probably connected with the biosynthesis of phenoxazinones, which are characteristic pigments of the genus Pycnoporus. Offprint requests to: B. Gross     

Influence of the mycelium growth conditions on the production of amylolytic,proteolytic and pectinolytic enzymes by Aspergillus niger C

Various nitrogen and carbon sources, as well as natural products, were examined as inducers of the production of amylases, proteases and pectinases by A. niger C. A. niger C grown on wheat bran extract medium provided culture supernatants with the highest enzymatic activities. Some culture conditions, e.g. pH, medium temperature and time period of cultivation, were optimalized to improve the growth and enzymes biosynthesis by A. niger C.     

Biosynthesis of cephamycin by resting cells of Streptomyces lactamdurans L 2/6

In order to investigate the nutritional conditions of cephamycin biosynthesis independently of the biomass growth process, the nutrient limited-resting cell system was used. A replacement medium eliminating cell multiplication was developed. The presence of Mg2+, carbon source and nitrogen source was necessary for cephamycin production by resting cells of Streptomyces lactamdurans L 2/6. Maximum antibiotic production was obtained when maltose, saccharose, and fructose were used as carbon source, and L-asparagine as nitrogen source. An inhibitory effect on the process was exerted by the calcium ions. There was no visible inhibition of cephamycin biosynthesis by inorganic phosphate ions in concentration up to 100 mM.     

Culture conditions for the production of amylolytic enzymes by a thermophilic Clostridium sp.

The growth of a thermophilic Clostridium sp. and the production of α-glucosidase, α-amylase and pullulanase were studied under anaerobic conditions using different carbon and nitrogen sources and varying pH values and temperatures. Growth and enzyme activities were highest with soybean meal as the nitrogen source. The optimum concentration was 2.5% [w/v] for the production of α-amylase as well as pullulanase and 2% [w/v] for α-glucosidase. The best carbon source proved to be soluble starch for α-amylase, and pullulanase and maltose for α-glucosidase. Growth and enzyme production reached their optimum at pH 6.5 to 7.0 and 70°C. Under these conditions, the enzyme activities followed exponential growth with maximum yields of α-glucosidase, α-amylase and pullulanase at 28, 36, and 44 h.     

Biosynthesis of extracellular low-molecular-mass papain and trypsin inhibitors by Streptomyces sp. 22: effect of cultural conditions

The production of extracellular inhibitors of papain and trypsin by Streptomyces sp. 22 was studied under different cultural conditions including complex and defined media, temperatures ranging from 18 °C to 37 °C and a variety of sole carbon and nitrogen sources. In complex nutritionally rich medium, maximal specific growth rates were obtained at 37 °C, whereas the highest specific production rates for both papain and trypsin inhibitors were registered at 18 °C. Studies on the effect of different carbon and nitrogen sources in defined media underline the importance of the nitrogen source as a strong regulator of the biosynthesis of both inhibitors. Enhanced formation of the inhibitory compounds occurred in the presence of casein. The dynamics of the formation of both inhibitors in defined media showed close association with growth. However, a partial separation of production phases for papain and trypsin inhibitors was observed in complex medium. The results imply differences in the regulation of biosynthesis of the two inhibitors.     

Enhancement of cephamycin C production using soybean oil as the sole carbon source

Vegetable oils were investigated to evaluate their potential to act as the sole carbon source for production of cephamycin C in shake and jar-fermentor cultures. Soybean oil was the best carbon source for cephamycin C production. Bioautography and HPLC analyses showed that cephamycin C was exclusively produced even when soybean oil was used as the sole cabon source. The optimal pH and initial concentration of soybean oil was 7.5 and 7 g/l, respectively. Both pH and the pH-control agent affected cephamycin C production, and among phosphoric acid, acetic acid and sulfuric acid, phosphoric acid was associated with the best production. Soybean oil was slowly consumed after the soluble nitrogen source was consumed. When the initial soybean oil concentration was 7 g/l, cephamycin C production was maximal, 2.0 g/l, which was twice as high as that from starch. The product yield from soybean oil was 4.7 times higher than that from starch. These results show that vegetable oils, which are cheaper than other carbon sources, could be used as the sole carbon source in the production of antibiotics. Correspondence to: M. Okabe     

Candida albicans cell wall lytic enzyme produced by Streptomyces thermodiastaticus

The production of lytic enzyme by Streptomyces thermodiastaticus was found to be affected by some growth conditions and nutritional factors. The highest enzyme production was obtained after 18 h of incubation at pH 5.5 and at 50 degrees C. The carbon source influenced the lytic enzyme production. A higher enzyme yield was obtained when Candida albicans cell wall (1 g/100 ml) was used as the sole carbon source. NaNO3 at 0.1 g/100 ml was the best nitrogen source for enzyme production. From all phosphorous sources, microelements, and growth factors tested, KH2PO4 (1 g/l), ZnSO4 (1 mg/I) and Tween 80 (0.1%), respectively, were found to favour the highest production of lytic enzymes by S. thermodiastaticus. The lytic enzymes mainly produced chitinolytic and proteolytic activities.