• Microbiology and Biotechnology Letters
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• v.37 no.3
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• pp.204-212
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• 2009

A strain producing ${\alpha}$-galactosidase and ${\beta}$-glucosidase was isolated from Kimchi. The isolated strain was identified as Weissella cibaria by 16S rDNA analysis and designated as Weissella cibaria K-M1-4. The enzyme activity of ${\alpha}$-galactosidase and ${\beta}$-glucosidase reached the maximum in the soy medium at $37^{\circ}C$ for 24 hr. The enzymes were purified by ethanol fractionation, DEAE sepharose fast flow, and sephacryl S-100HR column chromatography. ${\alpha}$-Galactosidase specific activity was shown by 576 Units/mg protein and the yield was 3.5% of the total activity of crude extracts. ${\beta}$-glucosidase specific activity was shown by 480 Units/mg protein and the yield was 2.9% of the total activity of crude extracts. The optimum temperature for ${\alpha}$-galactosidase was $60^{\circ}C$ and 43% of its original activity remained when it was treated at $80^{\circ}C$ for 30 min. For ${\alpha}$-galactosidase shows the optimum pH of 8.0 and is fairly stable between pH5.0 and pH9.0. The enzyme activity was increased in the presence of $Fe^{2+}$ and $Cu^{2+}$. The value of Km and Vmax for the enzyme were 0.98 mM and $1.81{\mu}$mole/min, respectively. The ${\beta}$-glucosidase has the optimum temperature of $50^{\circ}C$ and 46% of its original activity remained when it was treated at $80^{\circ}C$ for 30min. Its optimum pH of 7.0 and is fairly stable between pH5.0 and pH9.0. The enzyme activity was increased in the presence of $Fe^{2+},\;Co^{2+}$ and $Cu^{2+}$. The value of Km and Vmax for the enzyme were 1.24 mM and $6.81{\mu}$mole/min, respectively.

• Journal of Microbiology and Biotechnology
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• v.8 no.5
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• pp.484-489
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• 1998

Galactooligosaccharides (GalOS) were efficiently produced by partially purified $\beta$-galactosidase from the yeast strain Bullera singularis ATCC 24193. Ammonium sulfate precipitation and ultrafiltration methods were used to prepare the enzyme. The enzyme activity decreased at $50^{\circ}C$ and above. A maximum yield of 40% (w/w) GalOS, corresponding to 120 g of GalOS per liter, was obtained from 300 g per liter of lactose solution at $45^{\circ}C$, pH 3.7 when the lactose conversion was 70%. The yield of GalOS did not increase with increasing initial lactose concentration but the total amounts of GalOS did. Volumetric productivity was 4.8 g of GalOS per liter per hour. During this reaction, the by-products, glucose and galactose, were found to inhibit GalOS formation. Reaction products were found to be comprised of disaccharides and trisaccharides according to TLC and HPLC analyses. We propose the structure of the major product, a trisaccharide, to be ο-$\beta$-D-galactopyranosyl-(l-4)-ο-$\beta$-D-galactopyranosyl-(l-4)-$\beta$-D-glucose (4'-galactosyl lactose).

• Journal of Microbiology and Biotechnology
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• v.27 no.8
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• pp.1392-1400
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• 2017

Galactooligosaccharides (GOSs) are known to be selectively utilized by Bifidobacterium, which can bring about healthy changes of the composition of intestinal microflora. In this study, ${\beta}-GOS$ were synthesized using bifidobacterial ${\beta}-galactosidase$ (G1) purified from recombinant E. coli with a high GOS yield and with high productivity and enhanced bifidogenic activity. The purified recombinant G1 showed maximum production of ${\beta}-GOSs$ at pH 8.5 and $45^{\circ}C$. A matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis of the major peaks of the produced ${\beta}-GOSs$ showed MW of 527 and 689, indicating the synthesis of ${\beta}-GOSs$ at degrees of polymerization (DP) of 3 and DP4, respectively. The trisaccharides were identified as ${\beta}-{\text\tiny{D}}$-galactopyranosyl-($1{\rightarrow}4$)-O-${\beta}-{\text\tiny{D}}$-galactopyranosyl-($1{\rightarrow}4$)-O-${\beta}-{\text\tiny{D}}$-glucopyranose, and the tetrasaccharides were identified as ${\beta}-{\text\tiny{D}}$-galactopyranosyl-($1{\rightarrow}4$)-O-${\beta}-{\text\tiny{D}}$-galactopyranosyl-($1{\rightarrow}4$)-O-${\beta}-{\text\tiny{D}}$-galactopyranosyl-($1{\rightarrow}4$)-O-${\beta}-{\text\tiny{D}}$-glucopyranose. The maximal production yield of GOSs was as high as 25.3% (w/v) using purified recombinant ${\beta}-galactosidase$ and 36% (w/v) of lactose as a substrate at pH 8.5 and $45^{\circ}C$. After 140 min of the reaction under this condition, 268.3 g/l of GOSs was obtained. With regard to the prebiotic effect, all of the tested Bifidobacterium except for B. breve grew well in BHI medium containing ${\beta}-GOS$ as a sole carbon source, whereas lactobacilli and Streptococcus thermophilus scarcely grew in the same medium. Only Bacteroides fragilis, Clostridium ramosum, and Enterobacter cloacae among the 17 pathogens tested grew in BHI medium containing ${\beta}-GOS$ as a sole carbon source; the remaining pathogens did not grow in the same medium. Consequently, the ${\beta}-GOS$ are expected to contribute to the beneficial change of intestinal microbial flora.

• Microbiology and Biotechnology Letters
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• v.19 no.5
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• pp.456-463
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• 1991

$\beta$-Galactosidase of Bifidobacterium longum KCTC 3215 was studied on the production, purification, and characterization. Optimum conditions for the enzyme production were in the medium of 1.0% lactose as carbon source, initial pH 7.0 and in 17 hours of cultivation at $37^{\circ}C$. The enzyme was purified 9.25 folds by protamine sulfate precipitation, ammonium sulfate fractionation, DEAE-Sephadex A-50 ion exchange chromatography and Sephadex G-150 gel filtration. The maximal P-galactosidase activity was observed at pH 6.5 and at the temperature of $40^{\circ}C$ This enzyme was stable at pH 6.0-8.5. Metal ions such as $Ca^{2+} \;and \; Co^{2+}$, 2-mercaptoethanol, cysteine, and glutathione stimulated B-galactosidase activity. The enzyme activity was inhibited by addition of $Mg^{2+}, Fe^{2+}, Cs^{1+}, Li^{1+}$, DETA, galactose, and $\rho$-chloromercuribenzoic acid. The kinetics of o-nitrophenyl-$\beta$-D-galactopyranoside and lactose were $K_m$ = 1.66 mM, $V_{max}= 0.30 mM/min\cdot mg\cdot protein$ and $KK_m = 3.18 mM, \; V_{max}= 0.42 mM/min \cdot mg\cdot$ protein, respectively. The molecular weight of native enzyme was about 360, 000 dalton and the enzyme consisted of 2 identical subunits with a molecular weight of 180, 000.

• Asian-Australasian Journal of Animal Sciences
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• v.6 no.3
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• pp.373-375
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• 1993

Quantitative ${\beta}$-galactosidase estimation in the intestinal mucosal cells of calves with diarrhea under experimental conditions due to rotavirus were undertaken. A quantitative decrease of 40-70% in ${\beta}$-galactosidase activity was observed in proximal and middle segments of the small intestine of the infected calves, more so in the middle segments. The decrease in the distal part of the intestine, however, was lesser (5 to 30%). The decrease in the activity was more marked on the day 2 to 6 post infection indicating the degree of the damage of the villi of the small intestine.

• Applied Biological Chemistry
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• v.38 no.6
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• pp.507-511
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• 1995

In an attempt to improve the productivity of ${\beta}-galactosidase$ from Bacillus sp. A1, which was isolated from soil and has remarkably higher transgalactosylation activity than lactose hydrolysis activity, a chemical mutation procedure using N-methyl-N'-nitro-N-nitrosoguanidine followed by selection was conducted. The final selection, designated as Bacillus sp. A4442, turned out to show a substantially increased enzyme productivity. Catabolite repression by glucose and lactose requirement as an inducer for the enzyme biosynthesis, which were shown in the parent strain, was markedly diminished; instead it was found out that galactose acts as another inducer. Because pH of medium, one of the most important factors for cell growth as well as enzyme production, is closely related with the sugar concentration during culture, it was kept in the optimum range of $6.5{\sim}7.5$; for this the initial glucose concentration was adjusted to be 0.5% which was thereafter maintained by the controlled pumping-in of lactose using the pH-stat technique. By doing so, we were able to increase the productivity of ${\beta}-galactosidase$ with high transgalactosylation activity up to $44\;unit/m{\ell}-broth$.

• 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%.

• Applied Biological Chemistry
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• v.26 no.4
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• pp.217-221
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• 1983

$\beta$-Galactosidase($\beta$-D-galactoside galactohydrolase, E.C. 3. 2. 1. 23) from E. coli K-12 CSH 36 was immobilized on porous cellulose beads which were previously activated with tannin and p-benzoquinone. Their general properties and applicational possibities were investigated. The most effective, enzyme immobilization was obtained when tannin and p-benzoquinone, pH 11.0, were used together as activation reagents and a period of 6 hours of activation. The optimum pH of $\beta$-galactosidase was 5.5 for free enzyme and 6. 0 for the immobilized enzyme, the optimum temperatures for native and immobilized enzyme were both $50^{\circ}C$. Kms of native $\beta$-galactosidase and immobilized enzyme for ONPG(o-nitrophenyl galactopyranoside) were about $4.0{\times}10^(-4)M$ and $7.5{\times}10^(-4)M$, respectively. In the case of tannin : p-benzoquinone activated cellulose beads, the immobilized enzyme retained over 80% of the initial enzyme activity after 20 runs, which is very promising result far a possible industrial application.

• Applied Biological Chemistry
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• v.39 no.5
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• pp.333-337
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• 1996

For continuous production of galactooligosaccharides(GOS), $\beta-galactosidase$ with h1gh transgalactosylation activity from Bacillus sp. A 4442 was Immobilized onto $Diaion^{TM}$ HPA 75(styrene-divinylbenzene resin). The parameters influencing enzyme immobilization were scrutinized in order to maximize immobilization yield while minimizing enzyme inactivation. The optimum conditions turned out to be: Tris buffer concentration 30 mM, pH 8.0, contact time at room temperature 3 hr, and enzyme loading 25 mg protein/g resin. Both the thermal stability and the operational stability of immobilized enzyme were markedly enchanced by the treatment with 0.5% glutaraldehyde as a cross-linker. Under the experimental conditions established, the yield of ${\beta}-galactosidase$ immobilization was 40% or more and the activity of the immobilized enzyme ca. 200 U/g resin. When a packed-bed reactor was employed to continuously convert lactose to GOS, the specific production, which refers to as the amount of commercially valuable GOS produced by a unit amount of immobilized ${\beta}-galactosidase$, was found to be ca. 300 g GOS/g carrier.

• 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 $27^{\circ}C$.