• Microbiology and Biotechnology Letters
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• v.26 no.1
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• pp.40-44
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• 1998

A Penicillium strain which produces $eta$-galactosidase with high transgalactosylation activity, was isolated from soil and registered as Penicillium sp, KFCC 10888. When $eta$-galactosidase from Penicillium sp. KFCC 10855 reacted with 40% lactose, transgalactosylation ratio reached up to 70% at the 73% conversion of initial lactose. The biosynthesis of the enzyme in Penicillium sp. KFCC 10888 was not induced by lactose. The soybean meal was an effective component of the culture medium. The optimum pH and temperature for transgalactosylation were 4.0 and 55$^{\circ}C$, respectively. The production of galactooligosaccharides was in proportion to the initial lactose concentration. When the enzyme reacted with 40% lactose (pH 4.0) at 55$^{\circ}C$, the concentration of galactooligosaccharides increased up to 40% of total solid concentration.

• Journal of Microbiology and Biotechnology
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• v.8 no.4
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• pp.318-324
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• 1998

A ${\beta}$-galactosidase with high transgalactosylic activity was purified from a Bacillus species, registered as KFCC10855. The enzyme preparation showed a single protein band corresponding to a molecular mass of 150 kDa on SDS-PAGE and gave a single peak with the estimated molecular mass of 250 kDa on Sephacryl S-300 gel filtration, suggesting that the enzyme is a homodimeric protein. The amino acid and sugar analyses revealed that the enzyme is a glycoprotein, containing 19.2 weight percent of sugar moieties, and is much more abundant in hydrophilic amino acid residues than in hydrophobic residues, the mole ratio being about 2:1. The pI and optimum pH were determined to be 5.0 and 6.0, respectively. Having a temperature optimum at $70^{\circ}C$ for the hydrolysis of lactose, the enzyme showed good thermal stability. The activity of the enzyme preparation was markedly increased by the presence of exogenous Mg (II) and was decreased by the addition of EDTA. Among the metal ions examined, the most severely inhibitory effect was seen with Ag (I) and Hg (II). Further, results of protein modification by various chemical reagents implied that 1 cysteine, 1 histidine, and 2 methionine residues occur in certain critical sites of the enzyme, most likely including the active site. Enzyme kinetic parameters, measured for both hydrolysis and transgalactosylation of lactose, indicated that the enzyme has an excellent catalytic efficiency for formation of the transgalactosylic products in reaction mixtures containing high concentrations of the substrate.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.43 no.4
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• pp.373-379
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• 2017

We carried out the enzymatic synthesis of 1, 2-hexanediol galactoside (HD-gal) by transgalactosylation reaction using recombinant Escherichia coli ${\beta}-galactosidase$ (${\beta}-gal$). The amounts of ${\beta}-gal$ and 1, 2-hexanediol (HD), pH, and temperature, respectively, were first optimized (${\beta}-Gal$, 4.8 U/mL; HD, 75 mM; pH, 7.0; temperature, $37^{\circ}C$). Under these optimal conditions, about 96% HD was converted to HD-gal. When we investigated the water holding capacities (WHCs) of HD and HD-gal using pig epidermis in the concentrations of 84.4, 126.6, 168.8, 211.0 mM, WHC of HD-gal was superior to HD. In particular, at 168.8 mM HD and HD-gal, WHC of HD-gal showed about 20% greater than that of HD. However, it was observed that MIC values against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus of HD-gal were about three to ten times greater than those of HD, although MIC value of HD-gal against Enterococcus faecalis was almost the same as that of HD. Finally, it was concluded that the covalent bonding of a galactose molecule to HD (transgalactosylation) resulted in an increase in WHC of HD-gal and a decrease in anti-bacterial activity.

• Applied Biological Chemistry
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• v.41 no.7
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• pp.500-504
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• 1998

The enzymatic transgalactosylation of L-ascorbic acid was investigated to synthesize a chemically stable form of L-ascorbic acid by using commercially available ${\beta}-galactosidases$. Among various enzymes examined, Aspergillus oryzae ${\beta}-galactosidase$ was found to be formed the derivative of ascorbic acid in a high yield from ascorbic acid and lactose. The reaction product was isolated by ion exchange chromatography on a $Dowex\;1\;{\times}\;8$ (Cl - form) resin and Toyopearl HW-40S gel chromatography. The product was identified as $6-O-{\beta}-_D-galactopyranosyl-_L-ascorbic\;acid$ on the basis of various experimental results, viz., UV, IR, $^1H-NMR,\;^{13}C-NMR$ and mass spectral data.

• Applied Biological Chemistry
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• v.38 no.6
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• pp.502-506
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• 1995

A Bacillus strain which produces ${\beta}-galactosidase$ with high transgalactosylation activity, was isolated from soil and tentatively designated as Bacillus sp. A1. When ${\beta}-galactosidase$ from Bacillus sp. A1 reacted with 40% (w/w) lactose, transgalactosylation ratio reached up to 90% at the 70% conversion of the initial lactose. The biosynthesis of the enzyme in Bacillus sp. A1 required lactose as an inducer and was repressed by glucose. Observing that the addition of amino acids to culture medium resulted in enhancing, to a significant extent, both the growth and the enzyme production of the strain, yeast extract and commercially available hydrolysates of protein were examined for the suitability as amino acid source. As it turned out, SMP, an enzymatic hydrolysis product of soybean protein from Fuji Oil Co.(Japan), was the most suitable for optimization of the culture medium. When Bacillus sp. A1 was cultured in the presence of 0.5% SMP and 2% lactose, the enzyme activity increased up to $1.8\;U/m{\ell}-broth$.

• Journal of Life Science
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• v.26 no.5
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• pp.608-613
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• 2016

Recently, it has been reported that some preservatives used in cosmetics lead to skin problems. Among the many cosmetic ingredients, 1, 2-hexanediol (HD) is used as both a preservative and humectant. In order to develop safer ingredients, we studied the synthesis of 1, 2-hexanediol galactoside (HD-G) by a transgalactosylation reaction using β-galactosidase (β-gal)-containing recombinant Escherichia coli cells. The transgalactosylation reaction was carried out under high-lactose conditions for 24 hr. After 12 hr had elapsed, a new spot was identified by thin-layer chromatography (TLC) analysis, and we presumptively designated this new spot as HD-G. Then, we carried out the purification of the presumptive HD-G spot from the reaction mixture by using silica gel chromatography, and its mass was measured by electrospray ionization-mass spectrometry. The purified new spot on the chromatograph was identified a sodium adduct ion ([M+Na]⁺, m/z = 303.1423) of HD-G. In addition, when purified HD-G was hydrolyzed using commercially available E. coli β-gal, it was observed that a galactose molecule was released from HD-G. That is, it was demonstrated that HD-G is a galactoside derivative of HD. Finally, we confirmed that HD-G was enzymatically synthesized by E. coli β -gal as a new molecular entity. In the future, we plan to determine the minimum inhibitory concentrations of HD-G against different bacterial species. The cytotoxicity of HD-G against human skin cells will also be examined. It is expected hopefully that the galactosylation of HD would improve the functionality of HD-G.

• 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 $44\;unit/m{\ell}-broth$.

• Journal of the Korean Applied Science and Technology
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• v.36 no.2
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• pp.572-580
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• 2019

Recently, it has been reported that benzyl alcohol (BzOH) as an additive in cosmetics, food, and medicine lead to toxicity and allergy problem. Then, to circumvent this hurdle, we carried out the synthesis of benzyl alcohol galactoside (BzO-gal). Previously, it was confirmed that BzO-gal was synthesized by transgalactosylation reaction using Escherichia coli (E. coli) ${\beta}$-galactosidase (${\beta}-gal$). Meanwhile, in this study, two peaks of BzO-gal as sodium adduct ion (m/z=293.1004) and protonated ion (m/z=271.1180) were detected in the reaction mixture by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS). In addition, the amount of ${\beta}-gal$ and BzOH concentration, temperature, pH, and lactose concentration, respectively, were optimized (${\beta}-gal$, 0.75 U/mL; BzOH, 185 mM; temperature, $40^{\circ}C$, pH, 7.5; lactose, 350 g/l). Under these optimal conditions, 185 mM BzOH was converted into about 131 mM BzO-gal, in which the conversion yield was about 72%. In the future, BzO-gal will be applicable as a substitute for BzOH as a less toxic preservative for the cosmetic, pharmaceutical, and food industries, and we are planning to investigate the characteristics of BzO-gal as a preservative.

• KSBB Journal
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• v.11 no.3
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• pp.317-322
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• 1996

Galactooligosaccharlde (GOS) is a kind of functional oligosaccharides that can be used as a food ingredient and a cosmetic additive. In this paper, characteristics of GOS synthesis by cellulase, using lactose as a substrate, were investigated. Penicillium funiculosum cellulose was found to be the most efficient for GOS production among six cellulose tested. The optimum pH and temperature for GOS production were 5.0 and $50^{\circ}C$, respectively. There was an optimum ratio of lactose concentration to enzyme loading; the value was 10 (w/w). The reaction pattern of P. funiculosum cellulase is consistent with that of microbial ${\beta}$-galactosidase which shows transgalactosylation activity. Amounts of GOS produced from 20% (w/v) lactose after 6 h incubation at $50^{\circ}C$, were 23% (w/w) based on total saccharide in the reaction medium. The GOS % increased with initial lactose concentration in the range of 5 to 20%. The products mainly consisted of a trisaccharide and tetrasaccharide from HPLC and TLC analysis. Among enzymes involved in transgalactosylation reaction, high molecular weight fractions over 50,000 Da, presumably ${\beta}$-glucosldase, were considered to be responsible for GOS production. Using this cellulose, a direct synthesis of galactosyl g1ucoside including GOS could be readily achieved with lactose as a galactosyl donor.

• Food Science of Animal Resources
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• v.33 no.5
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• pp.565-571
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• 2013

An enzyme ${\beta}$-galactosidase or ${\beta}$-galactohydrolase [EC3.2.1.23], commonly called lactase, mediates galacto-oligosaccharide (GOS) synthesis under conditions of high substrate concentrations. Also, lactase hydrolyzes ${\beta}$($1{\rightarrow}4$) lactose into glucose and galactose, the latter is successively transferred to free lactose to make various oligosaccharides via transgalactosylation. GOS is non-digestible to human digestive enzymes and has been used as a functional prebiotics. Among the 24 lactic acid bacteria (LAB) strains used, Lactobacillus paracasei YSM0308 was selected based on its exhibition of the highest ${\beta}$-galactoside hydrolysis activity, and the crude lactase was prepared for examination of reaction conditions to affect the GOS synthesis. Lactase activity was measured with a spectrophotometer using ONPG (o-nitropheyl ${\beta}$-D-galactopyranoside) method. Lactase activity was not detected in the culture supernatant and was mostly present in the cell pellet after centrifugation. Activity of the crude lactase preparation ranges from102 to 1,053 units/mL, with the highest activity determined for L. paracasei YSM0308. Optimal conditions for GOS synthesis are as follows: concentration of whey powder, pH, temperature, and time were 30%, pH 6.5-7.0, $30^{\circ}C$, and 4 h, respectively. The final GOS concentration was 19.41% (w/v) by the crude YSM0308 lactase, which was obtained from strain YSM0308 grown in the 10% (w/v) reconstituted whey-based medium.