• Korean Journal of Microbiology
• /
• 제35권4호
• /
• pp.315-321
• /
• 1999

This study has been performed to deveope a yeast strain having high ${\alpha}$-amylase production ability using nuclear transfer method. Hybrids formed between the strains of Saccharomyces fiburigera KCTC 7393 and Saccharomyces cerevisiae KCTC 7049 (tyr-, ura-)were obtained by nuclear transfer technique. Nuclei isolated from the wild type S. fiburigera strain were transfered into auxotrophic mutants S. cerevisiae and selected the hybrids showing an increased starch degrading capability were selected (MN-16). This transformant grew best and produced maximal ${\alpha}$-amylase activity on the medium containing 2% (V/V) soluble starch. ${\alpha}$-Amylase from MN-16 was purified electrophoretically homogenety and its properties were investigated. The enzyme was purified about 10.6 fold with an overall yield 9.7% from the culture medium by ammonium sulfate fractionation. DEAE-Sephacel column chromatography, and Sephacryl S-200 column chromatography. The purified enzyme showed a single band on SDS-polyacrylamide gel electrophoresis. The molecular weight of the ${\alpha}$-amylase was estimated to be 53,000 daltons by SDS-PAGE and by gel permeation chromatography on Sephacryl S-200. The purified enzyme showed the maximum activity at pH 5.5 and 40${\circ}C$. The km value for soluble starch was 2.5㎎/㎖. The enzyme activity increased in the presence of $Ca^{2+}, Co^{2+}, EDTA, Mg^{2+}, Mn^{2+}, Zn^{2+}$, but inhibited by $Cu^{2+}, Fe^{2+}$, and $Ni^{2+}$

• Journal of Animal Science and Technology
• /
• 제63권6호
• /
• pp.1301-1313
• /
• 2021

This study investigated the effects of applying cellulase and starch on the fermentation characteristics and microbial communities of Napier grass silage after ensiling for 30 d. Three groups were studied: No additives (control); added cellulase (Group 1); and added cellulase and starch (Group 2). The results showed that the addition of cellulase and starch decreased the crude protein (CP), neutral detergent fiber (NDF), acid detergent fiber (ADF) and pH significantly (p < 0.05) and increased water-soluble carbohydrate (WSC) content (p < 0.05). The addition of additives in two treated groups exerted a positive effect on the lactic acid (LA) content, lactic acid bacteria (LAB) population, and lactic acid / acetic acid (LA/AA) ratio, even the changes were not significant (p > 0.05). Calculation of Flieg's scores indicated that cellulase application increased silage quality to some extent, while the application of cellulase and starch together significantly improved fermentation (p < 0.05). Compared with the control, both additive groups showed increased microbial diversity after ensiling with an abundance of favorable bacteria including Firmicutes and Weissella, and the bacteria including Proteobacteria, Bacteroidetes, Acinetobacter increased as well. For alpha diversity analysis, the combined application of cellulase and starch in Group 2 gave significant increases in all indices (p < 0.05). The study demonstrated that the application of cellulase and starch can increase the quality of Napier grass preserved as silage.

• Korean Journal of Microbiology
• /
• 제26권2호
• /
• pp.73-81
• /
• 1988

$\alpha$-Amylase (EC.3.2.1.1) of Neurospora crassa (ATCC9279) was cloned in E. coli HB101 using shotgun method, and the enzymes isolated from both N. crassa and E. coli were compared. Chromosomal DNA isolated from the spores of N. crassa was partially digested with PstI restriction endonuclease and rejoined to pBR322 which had been digested with the same enzyme. The resulting recombinant DNA were introduced into E. coli HB101 which had competancy by treating with $CaCl_{2}$. As the result, about 8000 colonies which showed tetracycline resistance were selected and two of the colonies which had 13.5Kb recombinant plasmid exhibit starch degrading activity on starch-containing plate when treated with D-cycloserine. $\alpha$-Amylases from both N.crassa and E. coli were isolated by using ammonium sulfate precipitation, DEAE-cellulose ion exchange column chromatography and Bio-Gel P150 gel foltration column. As the result, about 81.3 fold and 5.6 fold purifications in specific activities were obtained respectively, and specific activities of the gel filtrates were 6.1u/mg and 85u/mg respectively. The properties of both enzymes were compared and they showed quite the similar patterns in optimal temperature, optimal pH and had same molecular weight about 100,000 daltons on gel filtration method. Optimal temperatures for both enzymes were $70^{\circ}C$ and optimal pH were about 6 and 10.

• Mycobiology
• /
• 제39권1호
• /
• pp.33-39
• /
• 2011

Wild yeasts on the surface of various fruits including grapes were surveyed to obtain yeast strains suitable for fermenting a novel wine with higher alcohol content and supplemented with rice starch. We considered selected characteristics, such as tolerance to alcohol and osmotic pressure, capability of utilizing maltose, and starch hydrolysis. Among 637 putative yeast isolates, 115 strains exhibiting better growth in yeast-peptone-dextrose broth containing 30% dextrose, 7% alcohol, or 2% maltose were selected, as well as five ${\alpha}$-amylase producers. Nucleotide sequence analysis of the 26S rDNA gene classified the strains into 13 species belonging to five genera; Pichia anomala was the most prevalent (41.7%), followed by Wickerhamomyces anomalus (19.2%), P. guilliermondii (15%), Candida spp. (5.8%), Kodamaea ohmeri (2.5%), and Metschnikowia spp. (2.5%). All of the ${\alpha}$-amylase producers were Aureobasidium pullulans. Only one isolate (NK28) was identified as Saccharomyces cerevisiae. NK28 had all of the desired properties for the purpose of this study, except ${\alpha}$-amylase production, and fermented alcohol better than commercial wine yeasts.

• Journal of Pharmaceutical Investigation
• /
• 제20권4호
• /
• pp.209-215
• /
• 1990

The activities of s-amylase, ${\alpha}-amylase$ and protease of three combination products containing digestive enzymes, antacids and herbal drugs on the Korean market were estimated. The effects of antacids and herbal drugs on the activities of digestive enzymes were investigated. Starch-saccarifying activity of s-amylase, starch-dextrinizing activity of ${\alpha}-amylase$ and protein-peptic activity of protease were estimated by Somogy, Mc'Credy, and Casein-Folin method, respectivley. The optimal pH of s-amylase, ${\alpha}-amylase$ and protease were pH 5.0, 4.8 and 7.0, rcspectively. The digestive activities at optimal pH continued about eight hours. The digestive activities of individual enzymes were reduced to 40-90% by antacids and were affected somewhat positively or negatively by herbal drugs. Enzyme activities of the combination products were also affected by pH and reaction time.

• Journal of the Korean Society of Food Science and Nutrition
• /
• 제12권4호
• /
• pp.374-381
• /
• 1983

With a view of improving the quality of sardine sausage, the processing conditions of sardine sausage used raw sardine as materials and the effects of adding soybean protein and smoke flavor on the quality of product were investigated. Optimal amounts of additives in processing sardine sausage were 1.5% of salt, 1.5% of sugar, 0.2% of monosodium glutamate, 0.2% of white pepper, 0.2% of garlic powder, 0.2% of nutmeg, 0.4% of beef extract, 0.05% of food color solution(10% mixture solution of Red 40 and Yellow 5) and 0.1~0.2% of smoke flavor(Smok-EZ, Alpha Foods Co., Ltd.) based on washed sardine meat. The results showed that the benificial effects of adding corn starch(5%), ${\alpha}$-starch(2%), soybean protein(3%) to the washed sardine meat were exhibited in the improvement of texture and acceptability.

• KSBB Journal
• /
• 제19권2호
• /
• pp.164-167
• /
• 2004

Cyclodextrin glucanotransferase (EC 2.4.1.19, abbreviated as CGTase) is one of the most applied industrial enzymes that produces cyclodextrins from starch and related ${\alpha}$-1,4-glucans by intramolecular transglycosylation reaction upon Ca$\^$2＋/ dependent manner. The reaction of CLEC, ${\alpha}$-CGTases from Bacillus macerans with the soluble starch as a substrate reveals that the surfactants (SDS, N-octyl-${\beta}$-D-glucoside) significantly affect not only the overall products of CDs but also their selectivity. The surfactants (SDS, Lubrol PX) trigger the increase of ${\alpha}$-CD production, but Triton x-100 and Tween 80 suppress ${\alpha}$-CD specificity. Organic solvents (dimethyl sulfoxide, formamide, 2-methyl-2,4-pentandiol, and ethylene glycol) also cause changes of total product and product selectivity.

• Archives of Pharmacal Research
• /
• 제8권2호
• /
• pp.67-75
• /
• 1985

To find emylase inhibitors produced by microorganisms from soil, a strain which had a strong inhibitory activity against bacteria .alpha.-amylase was isolated from the soil smaple collected in Seoul. The morphological and physiological characteristics of this strain on several media and its utilization of carbon sources showed that it was one of Streptomyces specties according to the international Streptomyces Project method. The amylase inhibitor of this strain was purified by means of acetone precipitation, adsorption on Amberlite XAD-2, and column chromatography on Amberlite CG-50 and SP-Sephadex C-25. The inhibitor was stable at the pH range of 1-10 and at 100.deg.C for half an hour, and had inhibitory activities against other amylases such as salivary .alpha.-amylase, pancreatic .alpha.-amylase, fungal .alpha.-amylase and glucoamylase. The kinetic studies of the inhibitor showed that its inhibitory effect on starch hydrolysis by .alpha.-amylase was non-competitive.

• Microbiology and Biotechnology Letters
• /
• 제17권4호
• /
• pp.370-378
• /
• 1989

An Alkalophilic Bacillus circulans that can produce significant amount of cyclodextrin glucanotransferase (CGTase) was newly isolated from soil. The culture filtrate was successively purified by ($NH_4$)$_2$$SO_4$precipitation, DEAE-Sephadex column chromatography, and Sephadex G-100 column chromatography. The enzymatic properties, including molecular weight, optimal pH and temperature, stability, and kinetic parameters, were determined. The cyclodextrin synthesis reaction catalized by the purified CGTase was also studied. The sweet potato and corn starch were found to be the most suitable substrates with 60% conversion to cyclodextrin. The highest conversion was achieved at the CGTase concentration of 900-1,100 units/g of soluble starch. The purified CGTase could also catalize the transglycosylation on stevioside.

• Applied Biological Chemistry
• /
• 제33권1호
• /
• pp.79-86
• /
• 1990

Cyclodextrin hydrolase from Bacillus stearothermophilus KFCC 21203 was purified and the properties of the purified enzyme were investigated. The enzyme was purified 15 folds with 77 % recovery by ammonium sulfate fractionation, DEAE-cellulose chromatography, and Ultro AcA 34 gel filtration. The specific activity and the molecular weight of the enzyme were 1.30 units/mg protein and about 29,500, respectively, The maximum activity of the enzyme was shown at $55^{\circ}C$ and pH 5.5. However, stable temperature and pH were $40^{\circ}C$ and $5.0{\sim}8.0$, respectively. The Km value for ${\gamma}-cyclodextrin$ was $3.78{\times}10^{-3}$ M. The degradation activity of the enzyme was selectively high for ${\gamma}-cyclodextrin$, and very low for ${\beta}-cyclodextrin$, but not for ${\alpha}-cyclodextrin$. The decomposed products of ${\gamma}-cyclodextrin$ were mainly glucose and maltose, and a little mlatotriose. The activity of the enzyme was very high for amylose, potato starch, corn starch, amylopectin and maltooligomer, and relatively high for glycogen and dextrin. The decomposed products of them were mainly glucose and maltose.