• Food Science and Biotechnology
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• 제16권5호
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• pp.778-782
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• 2007

This study assessed the effects of the antimicrobial substances sulforaphane, grapefruit seed extracts (GSE), and reuterin on the expression of Salmonella HilA and InvF virulence gene using a LacZY assay (${\beta}$-galactosidase assay) with hilA:lacZY and invF:lacZY fusion strains of Salmonella typhimurium SL1344. Salmonella was grown for 8 hr at $37^{\circ}C$ in the presence of diluted antimicrobial substances ($2\;{\mu}g/mL$ sulforaphane, $20\{\mu}g/mL$ GSE, and 0.26 mM reuterin) at concentrations that did not inhibit the cellular growth of Salmonella. Sulforaphane inhibited the expression of HilA and InvF by 50-90 and 20-80%, respectively. GSE also inhibited the expression of both genes, but to a lesser degree. Among the 3 antimicrobial substances, reuterin showed the least inhibition, which was abolished after 3-4 hr. None of the antimicrobial substances inhibited the ${\beta}$-galactosidase enzyme activity of S. typhimurium. The assay used in this study represents a very sensitive method for screening bioactive substances that inhibit the expression of virulence genes in Salmonella.

• Journal of Microbiology and Biotechnology
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• 제1권3호
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• pp.151-156
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• 1991

In order to construct a yeast strain having high ethanol tolerance together with good lactose fermentation ability, the protoplast fusion using Saccharomyces cerevisiae STV 89 and Kluyveromyces fragilis CBS 397 was carried out. Auxotrophic mutants of K. fragilis were obtained as a selection marker by treatment of ethylmethane sulfonate. The best mutant for protoplast fusion was selected based on the capabilities of ${\beta}-galactosidase$ production and lactose fermentation. The protoplast fusion using polyethylene glycol and calcium chloride solution led to the fusion frequence of $3{\times}10^{-6}$ and a number of fusants were obtained. Among these fusants, a fusant F-3-19 showed the best results in terms of ethanol tolerance, ${\beta}-galactosidase$ activity and lactose fermentation. The performance of lactose fermentation and ethanol tolerance by this fusant were better than those of K. fragilis. Study on the ethanol tolerance having relation to fatty acid composition and intracellular ethanol concentration revealed that the fusant F-3-19 had a higher unsaturated fatty acids content and accumulated less amount of intracellular ethanol compared with a parent of K. fragilis.

• Asian-Australasian Journal of Animal Sciences
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• 제15권4호
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• pp.596-601
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• 2002

The present study was designed to examine the effect of ozone treatment in microencapsulated ${\beta}$-galactosidase on inactivation of the enzyme and sterilization of microorganism. The efficiency was the highest as 78.4% when the ratio of polyglycerol monostearate (PGMS) was 15:1. Activities of lactase remaining outside the capsule were affected by ozone treatment. With the increase of ozone concentration and duration of ozone treatment, the activity reduced significantly. In sensory aspect, with 2% microcapsule addition, no significant difference in sweetness was found compared with a market milk during 12 d storage. Above result indicated that the additional washing process of lactase was not necessary to inactivate the residual enzyme. In a subsequent study, the vegetative cells of microorganisms were completely killed with 10 ppm for 10 min treatment by ozone. The present study provides evidence that ozone treatment can be used as an inactivation and a sterilization process. In addition, these results suggest that acceptable milk products containing lactase microcapsules made by PGMS can be prepared with ozone treatment.

• Journal of Applied Biological Chemistry
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• 제53권1호
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• pp.1-7
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• 2010

xylA 프로모터는 대장균의 xylose 대사에 관여하는 xylose 오페론 상의 중요한 프로모터이다. 이 프로모터는 xylose에 의해 강하게 조절을 받는다고 알려져 있다. 이러한 특징은 새로운 발현 백터를 구축하는데 충분한 조건을 갖추고 있다고 생각된다. 본 연구에서는 이러한 xylose에 의해 유도 되는 발현벡터를 구축하기 위하여 600 bp의 xylA 프로모터를 증폭하여 pUC18의 AatII와 HindIII 사이에 삽입하여 pXA600을 구축하였다. 또한 조절단백질인 XylR의 영향을 조사하기 위하여 xylR 유전자를 삽입하여 pXAR600을 구축하였다. 발현의 강도를 측정하기 위하여 3,048 bp의 lacZ유전자를 xylA 프로모터의 하류에 연결하여 pXA600-lacZ와 pXAR600-lacZ를 구축하고 대장균 JM109에 형질전환시켰다. 구축된 pXA600-lacZ와 pXAR600-lacZ는 LB 배지에서 배양하였을 때 xylose 유도하에서 각각 1,641 unit와 2,304 unit의 $\beta$-galactosidase 활성을 보였으며, DM 배지상에서 배양했을 때 xylose 유도 시 각각 6,282 unit와 9,320 unit의 $\beta$-galactosidase 활성을 보였다. 또한 왜래 유전자의 발현 가능성을 확인하기 위하여 S. thermocyaneoviolaceus의 내열성 xylanase를 코딩하는 xynA 유전자를 실제로 구축된 pXA600과 pXAR600에서 발현을 확인하여 pXA600 및 pXAR600이 새로운 xylose 유도성 발현벡터로서의 사용 가능성을 확인하였다.

• 한국식품과학회지
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• 제32권1호
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• pp.218-224
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• 2000

분자량이 상이한 세 종류의 키토산의 항균활성을 E. coli O157 : H7, S. aureus 및 C. albicans 균주를 이용하여 측정, 분석하였다. E. coli O157 : H7와 S. aureus에 대해서는 분자량 10,000인 키토산이 가장 강한 항균활성을 보였으며 C. albicans에 대해서는 6량체의 키토산 올리고당이 가장 강한 활성을 나타내었다. 키토산 첨가농도는 E. coil O157 : H7와 S. aureus의 경우 0.1 mg/mL의 농도에서, C. albicans의 경우는 chitohexaose 1 mg/mL의 농도에서 항균활성이 가장 높았다. 모든 키토산 처리구에서 미생물 사멸 속도는 키토산 처리 1시간 이내에서 가장 높게 나타났으며 그 이후로는 점차 낮은 속도를 보였다. 사멸되었거나 파손된 미생물 세포로부터 유래되는 단백질, 핵산물질 및 $Ca^{2+}$ 량은 키토산 처리 시간 이내에 가장 많았으며 ${\beta}-galactosidase$ 활성도 같은 시간대에서 가장 빠른 속도로 중대되는 것으로 나타났다. S. aureus에 대한 세포막 손상 정도를 측정하여 본 결과 전체 미생물균체중 약 10%에 상당하는 균수가 막손상을 가져 왔다. 따라서, 키토산은 고유의 양이온성 성질을 이용하여 미생물의 세포벽과 세포막에 결합하여 그 결과로 세포 내 물질의 세포 외로의 유출내지는 세포막 대사의 저해 등의 효과를 나타냄으로써 항균활성을 나타내는 것으로 추측되었다.

• 한국축산식품학회지
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• 제33권5호
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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.

• Journal of Microbiology
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• 제37권3호
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• pp.175-179
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• 1999

To elucidate the role of vir-box in the expression of the virE gene, the vir-box was modified by site-directed mutagenesis and tested for ${\beta}$-galactosidase activities. A, C, T T, A, C substitutions at -62, -63, and -65 positions, destroying the 5'-region of the vir-box and A T at position -55, destroying the 3'-region of the vir-box respectively, showed only 17% promoter activity. When the vir-box was modified to contain perfect dyad symmetry structure (DSR) by the substitutions T, G A, T at -60 an d-61 positions, ${\beta}$-glactosidase activity increased 302%. These results indicate that the 5' and 3'-region of vir-box as well as the imperfect DSR of the vir-box itself may play a very important role in the regulation of virE gene expression.

• Journal of Microbiology and Biotechnology
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• 제15권4호
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• pp.689-693
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• 2005

Bacillus sp. JB-99, when grown in a chemically defined medium containing lactose as a carbon source, yielded 3,860 U/ml extracellular $\beta$-mannanase, which was high compared to other examined carbon sources. Among the nitrogen sources, yeast extract enhanced the enzyme activity. The enzyme production was growth-associated. The enzyme was optimally active at $65^{\circ}C$, pH 10, and had a half-life of 190 min at $65^{\circ}C$. N-Bromosuccinamide and $AgNO_3,\;CuSO_4$, and $HgCl_2$ strongly inhibited the enzyme, whereas $Ca^{2+}$ stimulated the enzyme activity. The $\alpha$-galactosidase enzyme production was not found in any of the enzyme assays.

• BMB Reports
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• 제33권2호
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• pp.166-171
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• 2000

Thioredoxin is a multifunctional protein that is ubiquitous in microorganisms, animals and plants. Previously, the expression of the Escherichia coli thioredoxin gene (trxA) was found to be negatively regulated by cAMP. In the present study, the effect of temperature on the expression of the E. coli trxA gene was investigated. In order to examine the temperature effect, the fusion plasmid pCL70 that harbors the E. coli trxA P1P2 promoter was used. The other two fusion plasmids, pJH3 and pMH521 that were constructed in different vectors which harbor the E. coli trxA P2 promoter, were also used. When the E. coli strain MC1061/pCL70 was grown in a rich medium at $25^{\circ}C$, $34^{\circ}C$ and $42^{\circ}C$, the cells grown at $42^{\circ}C$ gave the highest $\beta$-galactosidase activity. The E. coli MC1061/pJH3 and MC1061/pMG521 cells showed increased $\beta$-galactosidase activity after the shift of the culture temperature to $42^{\circ}C$. The wild-type trxA gene of the E. coli MC1061 cells produced much higher thioredoxin activity at the higher temperature. These results support the conclusion that the E. coli trxA gene is regulated in a temperature-dependent manner. Especially the expression from its P2 promoter appeared to be sensitive to temperature.

• 생명과학회지
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• 제15권2호
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• pp.215-219
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• 2005

감궁탕의 돌연변이원성을 유무를 알아보기 위해 Salmonella typhimurium TA 98 및 TA100을 이용한 돌연변이 원성 실험에서도 감궁탕은 어느 균에서도 돌연변이원성을 나타내지 않았으며, 이는 S-9 mixture 의해 감궁탕이 대사가 된 후에도 이와 유사한 경향을 나타내었다. 또한, SOS umu test의 경우에서도 $\beta-galactosidase$활성에는 별다른 영향을 미치지 않는 것으로 보아 감궁탕은 돌연변이원성을 일으키지 않는 것으로 판정되었으며 S-9 mixture처리에 의한 대사 후에도 이와 유사한 실험결과가 나타났다. 따라서 감궁탕은 그 자체 및 대사 후에도 DNA에 별다른 영향을 미치지 못하는 비교적 안전한 생약처방으로 여겨진다.