• Journal of Life Science
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• v.9 no.1
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• pp.26-34
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• 1999

A thermostable pullulanase has been isolated and purified from Thermus caldophilus GK-24 to a homogeneity by gel-filtration and ion-exchange chromatography. The specific activity of the purified enzyme was 431-fold increase from the crude culture broth with a recovery of 11.4%. The purified enzyme showed $M_{r}$ of 65 kDa on denaturated and natural conditions. The pI of the enzyme was 6.1 and Schiff staining was negative, suggesting that the enzyme is not a glycoprotein. The enzyme was most active at pH 5.5. The activity was maximal at $75^{\cire}C$ and stable up to $95^{\cire}C$ for 30 min at pH 5.5. The enzyme was stable to incubation from pH 3.5 to pH 8.0 at $4^{\cire}C$ for 24hr. The presence of pullulan protected the enzyme from heat inactivation, the extent depending upon the substrate concentration. The activity of the enzyme was simulated by $Mn^{2+}$ ion, }$Ni^{2+}$, $Ca^{2+}$, $Co^{2+}$ ions. The enzyme hydrolyzed the ${\alpha}$-1,6-linkages of amylopectin, glycogens, ${\alpha}$, ${\beta}$-limited dextrin, and pullulan. The enzyme caused the complete hydrolysis of pullulan to maltotriose and the activity was inhibited by $\alpha$, $\beta$, or $\gamma$-cyclodextrins. The $NH_{2}$-terminal amino acid sequence [(Ala-Pro-Gln-(Asp of Tyr)-Asn-Leu-Leu-Xaa-ILe-Gly-Ala(Ser)] was compared with known sequences of various sources and that was compared with known sequences of various sources and that was different from those of bacterial and plant enzymes, suggesting that the enzymes are structurally different.

• The Korean Journal of Food And Nutrition
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• v.10 no.3
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• pp.309-313
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• 1997

Bench scale Sikhyes were produced from rice and glutinous rice and limit dextrins in rice Sikhye and glutinous rice Sikhye were purified by ethanol precipitation and Biogel P-2 gel chromatography and FPLC on Superose 12 column and analyzed. The purified limit dextrin in rice Sikhye and glutinous rice Sikhye showed bot signal of $\alpha$-1,4- and $\alpha$-1,6-glucosidic linkage with its estimation ratio of 4.5:1 and 5.9:1, respectively, by 1H-NMR analysis. Limit dextrins were hydrolyzed by pullulanase. The enzyme hydrolysis products contained maltose, maltotriose, maltotetraose, maltopentaose and matohexaose. These results suggest that limit dextrins were composed of these maltoolgosaccharide series with $\alpha$-1,6-glucosidic bond.

• The Korean Journal of Food And Nutrition
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• v.10 no.1
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• pp.82-86
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• 1997

A Korean traditional sweet rice drink "Sikye" was produced from the raw material of 20% of rice and 4% malt supplemented with 2l of tap water, by incubating the mixture at 6$0^{\circ}C$ for 7 hours. The product was found to contain 11.01% of maltose, 5.31% of isomaltooligosaccharides, 1.75% of maltotriose and 0.28% of glucose. Maltose, maltotriose and isomaltooligosaccharides in Sikye were seperated by ethanol (3 volume) precipitation repeated three times, followed by gel chromatography of Toyopearl HW-40S. 1H-NMR analysis revealed that the products of G2 and G3 size had only $\alpha$-1, 4-glucosidic linkage. but isomaltooligosaccharides showed both signal of $\alpha$-1, 4 and $\alpha$-1, 6-glucosidic linkage with its estimation ratio of 5:1. Isomaltooligosaccharides were hydrolyzed to produce maltooligosaccharide series from maltose to maltohexaose by pullulanase. These results, suggest that isomaltooligosaccharides were constructed by maltohexaose main chain with maltose or maltotriose and maltotetraose side chain.ide chain.

• Journal of Microbiology and Biotechnology
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• v.26 no.11
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• pp.1933-1942
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• 2016

A novel ${\alpha}-glucoside$ hydrolase (named PspAG97A) from glycoside hydrolase family 97 (GH97) was cloned from the deep-sea bacterium Pseudoalteromonas sp. K8, which was screened from the sediment of Kongsfjorden. Sequence analysis showed that PspAG97A belonged to GH97, and shared 41% sequence identity with the characterized ${\alpha}-glucoside$ BtGH97a. PspAG97A possessed three key catalytically related glutamate residues. Mutation of the glutamate residues indicated that PspAG97A belonged to the inverting subfamily of GH97. PspAG97A showed significant reversibility against changes in salt concentration. It exhibited halophilic ability and improved thermostability in NaCl solution, with maximal activity at 1.0 M NaCl/KCl, and retained more than 80% activity at NaCl concentrations ranging from 0.8 to 2.0 M for over 50 h. Furthermore, PspAG97A hydrolyzed not only ${\alpha}-1,4-glucosidic$ linkage, but also ${\alpha}-1,6-$ and ${\alpha}-1,2-glucosidic$ linkages. Interestingly, PspAG97A possessed high catalytic efficiency for long-chain substrates with ${\alpha}-1,6-linkage$. These characteristics are clearly different from other known ${\alpha}-glucoside$ hydrolases in GH97, implying that PspAG97A is a unique ${\alpha}-glucoside$ hydrolase of GH97.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.1-7
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• 1989

To enhance the capability of starch fermentation of the transformant TSD-14, the heat treated protoplasts of TSD-14 were fused with the protoplasts of C. tropicalis (lys$^-$) in the presence of 30% (w/ v) PEG and 20 mM CaC1$_2$. Fusants were selected by nutritional complementation on minium medium and the fusion frequency was 4.4$\times$10$^{-5}$. All fusants tested were possessed of complemented traits concerning carbon compound assimilation, and the cell volumes of the fusants were approximately 1.5 times larger than the parental strains. The fusants were genetically very stable, and were able to hydrolyze alpha 1,4-glucosidic linkage as well as alpha 1,6-linkage of starch contrary to one of parents TSD-14, The most promising fusant FSC-14-75 produced 8.7% (v/v) of ethanol from 15% liquefied potato starch medium, but the result was enhanced to 9.3% (v/v) by addition of 0.3% peptone. The corresponding fermentation efficiency was 86.0%.

• The Korean Journal of Food And Nutrition
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• v.10 no.3
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• pp.370-374
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• 1997

Commercial Sikhye was fermented by Saccharomyces cerevisiae for 10 day at 29$^{\circ}C$. Sucrose was hydrolyzed into glucose and fructose by invertase from the yeast, and the glucose and fructose were converted into ethanol by the yeast. Commercial Sikhye wine was found to contain 6.6% of ethanol, 0.32$\mu$mol/ml of amino acid, 226$\mu\textrm{g}$/ml of protein, and 2.5ml 0.1N NaOH of acidity, respectively, and its pH was 3.21. Limit dextrin in commercial rice Sikhye wine showed both signal of $\alpha$-1, 4- and $\alpha$-1, 6- glucosidic linkage with its estimation ratio of 25:1 by 1H-NMR analysis. The taste of rice Sikhye wine was similar that of wine.

• The Korean Journal of Food And Nutrition
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• v.10 no.3
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• pp.360-364
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• 1997

Rice Sikhye was fermented by Saccharomyces cerevisiae for 10 day at 29$^{\circ}C$. Fermentable sugars such as glucose, maltose and maltotriose in rice Sikhye were converted into ethanol by the yeast, but limit dextrin was remained after the fermentation. Rice Sikhye wine was found to contain 5.3% of limit dextrin, 6.5% of ethanol, 2.9 $\mu$mol/ml of amino acid, 457$\mu\textrm{g}$/ml of protein, and the acidity of the Sikhye showed 3.1, respectively, and its pH was 3.67. Limit dextrin in rice Sikhye wine showed both signal of $\alpha$-1,4- and $\alpha$-1,6-glucosidic linkage wit its estimation ratio of 5.6:1 by 1H-NMR analysis. The taste of rice Sikhye wine was similar that of wine.

• YAKHAK HOEJI
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• v.32 no.1
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• pp.86-94
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• 1988

Some rheological properties of subfractions for dextran in the molecular weights range from $3{\times}10^4$ to $2{\times}10^6$ was investigated at room temperature. The dependence of the viscosity on concentration, shear rate, pH & ionic strength, temperature and solvent effect was observed. From the experimental data the Mark-Houwink viscosity equation in water at $25^{\circ}C$ was determined for samples having the molecular weight ranging from $3{\times}10^4$ to $2{\times}10^6$ as$[{\eta}]=3.1{\times}10^{-3}\;Mw^{0.39}(in\;dl/g)$. The intrinsic stiffness of the dextran backbone was estimated by evaluating the 'characteristic ratio' $C_{\infty}$, which is below the 0.082. In the concentrated region, the viscosity was decreased with increasing shear rate and was exponentially decreased with raising temperature, the viscosity showed the maximum value at neutral condition. From the experimental data, it was concluded that dextran chain, linked by the ${\alpha}-1$, 6-glucosidic linkage, behaves like a flexible random coil chain in aqueous solution, dextran solutions were pseudoplastic power law fluids among the empirical models of non-Newtonian behavior. Urea was an active reagent which increases the viscosity and swells dextran while pyrididine and glycerol were inactive reagents. Also, it could be estimated that the formation of gel structure is promote to the neutral state, the molecular weight larger than $2{\times}10^5$, when electrolytic concentration is IN and Ureas is use to solvent.

• The Korean Journal of Food And Nutrition
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• v.10 no.4
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• pp.453-457
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• 1997

Sugars in Korean and Japanese pumpkin were studied. The sugars in pumpkin were crushed and extracted by boiling for 30 min. Korean pumpkin was found to contain 0.41% of sucrose, 0.54% of fructose, 0.61% of glucose and 0.68% of starch. Japanese pumpkin was found to contain 2.60% of sucrose, 2.76% of fructose, 1.91% of glucose and 1.22% of starch. No other mono- and oligosaccharides were detected in the test of TLC and HPLC. Starch in Japanese pumpkin showed only signal of $\alpha$-1,4-glucosidic linkage by proton NMR analysis, and showed 86% of absorbance by iodine reaction compared with amylose(DP 117). These results indicated that starch in Japanese pumpkin is composed by only amylose. Pectin contents of Korean and Japanese pumpkin sowed 6.29% and 2.67%, respectively, as galacturonic acid by carbazole analysis.

• The Korean Journal of Mycology
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• v.27 no.1 s.88
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• pp.76-81
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• 1999

Exo-polysaccharide obtained from the submerged cultivation of Ganoderma lucidum mycelium was fractionated. The structural analysis of the acidic exo-polysaccharide fraction (BWS-DA-GI), showing high antitumor activity, was carried out and compared to the mycelial acidic fraction (MWS-DA-GI). The major sugar constituents of the fraction of BWS-DA-GI were glucose, galactose and mannose in the molar ratio of 2.5 : 2.1 : 2.5. The minor components in this fraction were xylose and fucose. While the major sugar constituents of the mycelial acidic fraction of MWS-DA-GI were galactose, fucose, mannose and glucose. The trace components in this fraction was xylose. From the results of periodate oxidation, Smith degradation, affinity chromatography and methylation analysis, the chemical structures of the two fractions, BWS-DA-GI and MWS-DA-GI were both determined as ${\beta}-1,3$ glucans. It was also estimated that BWS-DA-GI had a $1{\rightarrow}6$ glucosidic linkage and MWS DA-GI had $1{\rightarrow}4$ and $1{\rightarrow}6$ glucosidic linkages. The molecular weights of these fractions, MWS-DA-GI and MWSDA-GI were estimated as $1.2{\times}10^6\;and\;1.0{\times}10^6$ dalton, respectively.