• Korean Journal of Food Science and Technology
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• v.29 no.2
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• pp.376-378
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• 1997

In an attempt to increase the conversion of lactose to galactoligosaccharides, two types of ${\beta}-galactosidases$ originated from Thermus caldophilus and Bacillus sp. A4442 reacted with 60% (w/w) lactose consecutively. Concentration of galactooligosaccharides reached up to 60% at the 85% conversion of the initial lactose maintaining transgalactosylation ratio ca. 90%.

• Microbiology and Biotechnology Letters
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• v.31 no.1
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• pp.57-62
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• 2003

A bacteria strain producing extracellular $\beta$-mannanase was isolated from soil and was identified as Bacillus subtilis by 16S rRNA sequence comparison and biochemical determinations. The optimum pH and temperature for the $\beta$-mannanase activity were 5.0 and 5.5$^{\circ}C$, respectively. The zymogram technique revealed a single protein band exhibiting $\beta$-mannanase activity from the culture supernatant. The molecular mass of the enzyme was estimated at approximately 130 kDa. The addition of 0.5% lactose or 0.5% locust bean gum to the LB medium caused to Increase significantly the $\beta$-mannanase productivity from Bacillus subtilis JS-1. The cells grown on LB medium supplemented with lactose produced maximal enzyme activity at the stationary phase. In contrast to this, the $\beta$-mannanase was induced at the logarithmic phase from the cells grown on LB medium supplemented with locust bean gum. The discrepancy in induction times suggests that $\beta$-mannanase was induced by different induction mechanisms depending on the carbon sources in Bacillus subtilis JS-1 .

• Journal of Microbiology and Biotechnology
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• v.21 no.9
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• pp.972-978
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• 2011

In this study, we investigated the performance of an immobilized ${\beta}$-galactosidase inclusion bodies-containing Escherichia coli cell reactor, where the cells were immobilized in alginate beads, which were then used in repeated-batch operations for the hydrolysis of o-nitrophenyl-${\beta}$-D-galactoside or lactose over the long-term. In particular, in the Tris buffer system, disintegration of the alginate beads was not observed during the operation, which was observed for the phosphate buffer system. The o-nitrophenyl-${\beta}$-D-galactoside hydrolysis was operated successfully up to about 80 h, and the runs were successfully repeated at least eight times. In addition, hydrolysis of lactose was successfully carried out up to 240 h. Using Western blotting analyses, it was verified that the ${\beta}$-galactosidase inclusion bodies were sustained in the alginate beads during the repeated-batch operations. Consequently, we experimentally verified that ${\beta}$-galactosidase inclusion bodies-containing Escherichia coli cells could be used in a repeated-batch reactor as a biocatalyst for the hydrolysis of o-nitrophenyl-${\beta}$-D-galactoside or lactose. It is probable that this approach can be applied to enzymatic synthesis reactions for other biotechnology applications, particularly reactions that require long-term and stable operation.

• Microbiology and Biotechnology Letters
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• v.24 no.4
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• pp.493-499
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• 1996

Lactan gum produced by Rahnella aquatilis is a high viscous, anionic polysaccharide and has shear thinning behaviour. Lactan gum yield and cencentration was greater on disaccharide such as lactose and sucrose than on monosaccharides such as glucose and galactose. When initial carbon source concentration was 45g/l of sucrose of lactose, the microorgnisms produced 28 g/l and 27 g/l of lactan, respectively with a yield more than 60%. $\beta$-Galactosidase, hydrolyzing lactose into galactose and glucose, was induced by lactose or galactose. When initial corbon source was 45 g/l of mixed carbon I (glucose:galactose=1:1), lactan gum concentaration was higher than that from 45 g/l of monosaccharide (glucose pf galactose) but was similar to the result from 45 g/l of lactose. Therefore, lactose hydrolysis reaction by $\beta$-galactosidase does not seem to be a rate determining step in lactan gum biosynthesis. When initial carbon source was 45 g/l of mixed carbon II (glucose:fructose=1:1). total carbon source consumption rate was slower than that from sucrose, but glucose consumption rate was faster than that from fructose.

• Applied Biological Chemistry
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• v.23 no.4
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• pp.218-221
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• 1980

From S. lactis $KB_{21}$, stabilized strain by intergration of the lactose plasmid into the host chromosome, several lactose constitutive mutants were obtained by UV irradiation and spontaneous mutation. The rapid growth of the mutants in the medium containing lactobionate confirmed their constitutive nature. The mutants synthesized $phospho-{\beta}-galactosidase$ with an activity of 1.7 to 3.4 times that of the parent.

• Journal of Microbiology and Biotechnology
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• v.5 no.1
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• pp.6-13
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• 1995

A gene coding for the $\beta$-galactosidase (lactase) of Kluyveromyces tragilis UCD 55-55 was isolated by complementation in Escherichia coli YMC9. From the plasmid library made from Sau3A-digested chromosomal DNA, one positive clone was selected. The cloned gene for $\beta$-galactosidase was on 7.3 kilobase pair DNA fragment, and a slightly low level of $\beta$-galactosidase enzyme activity was detecied in E. coli. It was also confirmed that the cloned gene comes from K. tragilis by DNA-DNA hybridization and immunochemical blotting experiments. In order to construct a new yeast strain having the metabolic ability for lactose, the cloned gene for K. tragilis $\beta$-galactosidase was inserted in yeast vector YEp24 and YRp17, and transformed into Saccharomyces cerevisiae YNN27 and Ml-2B. The yeast transformants showed the nearly the same $\beta$-galactosidase productivity as level of K. tragilis when uninduced, but these could not utilize lactose as a sole carbon source, presumably due to the lack of lactose transport system. Nevertheless, a slightly higher ethanol productivity was achieved by these transformants than S. cerevisiae or K. tragilis, in the medium containing glucose and lactose.

• Korean Journal of Food Science and Technology
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• v.12 no.1
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• pp.45-52
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• 1980

The nylon (poly 6, 10) microcapsules containing ${\beta}-galactosidase$ were obtained by the interfacial polymerization of 1, 6-diaminohexane and sebacoyl chloride with ${\beta}-galactosidase$ from Escherichia coli. They were generally spherical and had a mean diameter of $80{\mu}$ with 45 % of the activity recovery. In particular, there was no transport hamper of lactose through the membrane of microcapsules. The characteristics of the microencapsulated enzyme were similar to those of soluble enzyme optimal pHs, $7.0{\sim}7.2$ for the soluble and $7.3{\sim}7.5$ for the microencapsulated ; optimal temperatures, $50^{\circ}C$ for both ; apparent $K_m,\;3.33{\times}10^{-4}(on ONPG),$ $2.86{\times}10^{-3}$ M(on lactose) for the soluble and $5.28{\times}10^{-4}$ (on ONPG), $4.25{\times}10^{-3}$ M (on lactose) for the microencapsulated ; activation energies, 8.94 for the soluble and 9.78 Kcal/mole for the microencapsulated enzyme. Using this microencapsulated ${\beta}-galactosidase$, hydrolyses of lactose and milk lactose were carried out and 80 % of 5 % lactose solution and 70 % of lactose in skim milk were hydrolyzed in 40 hr at $27^{\circ}C$. The reusability and operational stability showed that the remaining activity was 50 % of the original activity after 5 runs and 120 hr of total operating time at $27^{\circ}C$.

• Bulletin of the Korean Chemical Society
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• v.26 no.12
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• pp.2001-2006
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• 2005

Glycoproteins and glycolipids play key roles in intracellular reactions between cells and their environments at the membrane surface. For better understanding of the nature of these events, it is necessary to know threedimensional structures of those carbohydrates, involved in them. Since carbohydrates contain many hydroxyl groups which can serve both as hydrogen bond donors and acceptors, hydrogen bond is an important factor stabilizing the structure of carbohydrate. DMSO is an aprotic solvent frequently used for the study of carbohydrates because it gives detailed insight into the intramolecular hydrogen bond network. In this study, conformational properties and the hydrogen bonds in $\beta$-lactose in DMSO are investigated by NMR spectroscopy and molecular dynamics simulations. NOEs, temperature coefficients, deuterium isotope effect, and molecular dynamics simulations proved that there is a strong intramolecular hydrogen bond between O3 and HO2' in $\beta$-lactose and also OH3 in $\beta$-lactose may form an intermolecular hydrogen bond with DMSO.

• Journal of Microbiology and Biotechnology
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• v.21 no.11
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• pp.1151-1158
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• 2011

In this study, a galactooligosaccharide (GOS) was synthesized using active ${\beta}$-galactosidase (${\beta}$-gal) inclusion bodies (IBs)-containing Escherichia coli (E. coli) cells. Analysis by MALDI-TOF (matrix-assisted laser desorption/ionization-time of flight) mass spectrometry revealed that a trisaccharide was the major constituent of the synthesized GOS mixture. Additionally, the optimal pH, lactose concentration, amounts of E. coli ${\beta}$-gal IBs, and temperature for GOS synthesis were 7.5, 500 g/l, 3.2 U/ml, and $37^{\circ}C$, respectively. The total GOS yield from 500 g/l of lactose under these optimal conditions was about 32%, which corresponded to 160.4 g/l of GOS. Western blot analyses revealed that ${\beta}$-gal IBs were gradually destroyed during the reaction. In addition, when both the reaction mixture and E. coli ${\beta}$-gal hydrolysate were analyzed by high-performance thin-layer chromatography (HP-TLC), the trisaccharide was determined to be galactosyl lactose, indicating that a galactose moiety was most likely transferred to a lactose molecule during GOS synthesis. This GOS synthesis system might be useful for the synthesis of galactosylated drugs, which have recently received significant attention owing to the ability of the galactose molecules to improve the drugs solubility while decreasing their toxicity. ${\beta}$-Gal IB utilization is potentially a more convenient and economic approach to enzymatic GOS synthesis, since no enzyme purification steps after the transgalactosylation reaction would be required.

• Journal of Microbiology and Biotechnology
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• v.14 no.3
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• pp.584-592
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• 2004

A thermostable $\beta$-glycosidase gene, tfi $\beta$-gly, was cloned from the genomic library of Thermus filiformis Wai33 A1. ifi $\beta$-gly consists of 1,296 bp nucleotide sequence and encodes a polypeptide of 431 amino acids. It shares a strong amino acid sequence similarity with the $\beta$-glycosidases from other Thermus spp. belonging to the glycosyl hydrolase family 1. In the present study, the enzyme was overexpressed in Escherichia coli BL21 (DE3) using the pET21b(+) vector system. The recombinant enzyme was purified to homogeneity by heat treatment and a $Ni^{2+}$-affinity chromatography. Polyacrylamide gel electrophoresis (PAGE) showed that the recombinant Tfi $\beta$-glycosidase was a monomeric form with molecular mass of 49 kDa. The temperature and pH range for optimal activity of the purified enzyme were 80- $90^{\circ}C$ and 5.0-6.0, respectively. Ninety-three percent of the enzyme activity was remained at $70^{\circ}C$ after 12 h, and its half-life at $80^{\circ}C$ was 6 h, indicating that Tfi $\beta$-glycosidase is highly thermostable. Based on its K_m$, or $K_{cat}K_m$, ratio, Tfi $\beta$-glycosidase appeared to have higher affinity for $\beta$-D-glucoside than for $\beta$-D-galactoside, however, $K_{cat} for \beta$-D-galactoside was much higher than that for $\beta$-D-glucoside. The activity for lactose hydrolysis was proportionally increased at $70^{\circ}C$ and pH 7.0 without substrate inhibition until reaching 250 mM lactose concentration. The specific activity of Tfi TEX>$\beta$-glycosidase on 138 mM lactose at $70{^\circ}C$ and pH 7.0 was 134.9 U/mg. Consequently, this newly cloned enzyme appears to have a valuable advantage of conducting biotechnological processes at elevated temperature during milk pasteurization in the production of low-lactose milk.