• Journal of Microbiology and Biotechnology
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• v.20 no.4
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• pp.803-808
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• 2010

Poly-${\gamma}$-glutamic acid (${\gamma}$-PGA) is a natural edible polypeptide in which glutamate is polymerized via ${\gamma}$-amide linkages. First, we assessed the eye irritancy potential of ${\gamma}$-PGA in rabbits. Additionally, we studied the effects of ${\gamma}$-PGA on corneal wound healing, due to the anti-inflammatory properties and water retaining abilities of ${\gamma}$-PGA. In this study, the effects of ${\gamma}$-PGA on corneal wound healing after an alkali burn were evaluated. Thirty eyes wounded by alkali burning in 30 white rabbits were divided into three groups: group A was treated with 0.1% 5,000 kDa ${\gamma}$-PGA for 2 days; group B was treated with 0.1% hyaluronic acid; and group C was not treated, as a control. The area of corneal epithelial defect was examined at 12, 24, 30, 36, 42, and 48 h after corneal alkali wounding to determine initial wound healing. We found that ${\gamma}$-PGA promoted corneal wound healing, compared with controls, and showed similar effects to hyaluronic acid. These results indicate that ${\gamma}$-PGA stimulates corneal wound healing by an anti-inflammatory effect and enhancing cell migration and cell proliferation. ${\gamma}$-PGA is a promising biomaterial that may be a substitute for hyaluronic acid in corneal wound healing treatment.

• Journal of Microbiology and Biotechnology
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• v.24 no.2
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• pp.197-208
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• 2014

A novel protease gene from Bacillus gibsonii, aprBG, was cloned, expressed in B. subtilis, and characterized. High-level expression of aprBG was achieved in the recombinant strain when a junction was present between the promoter and the target gene. The purified recombinant enzyme exhibited similar N-terminal sequences and catalytic properties to the native enzyme, including high affinity and hydrolytic efficiency toward various substrates and a superior performance when exposed to various metal ions, surfactants, oxidants, and commercial detergents. AprBG was remarkably stable in 50% organic solvents and retained 100% activity and stability in 0-4 M NaCl, which is better than the characteristics of previously reported proteases. AprBG was most closely related to the high-alkaline proteases of the subtilisin family with a 57-68% identity. The secretion and maturation mechanism of AprBG was dependent on the enzyme activity, as analyzed by site-directed mutagenesis. Thus, when taken together, the results revealed that the halo-solvent-tolerant protease AprBG displays significant activity and stability under various extreme conditions, indicating its potential for use in many biotechnology applications.

• Journal of The Korean Society of Grassland and Forage Science
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• v.36 no.3
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• pp.191-198
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• 2016

This study evaluated the effects of TMC (trace mineral-fortified microbial culture) supplementation on growth performance, carcass characteristics, and meat quality parameters of Hanwoo steers during the last 4 months of finishing period. The TMC was a combination of 0.4% trace minerals, 20.0% Na-bentonite, and 79.6% feedstuffs, which was inoculated with a mixed microbial culture (Enterobacter ludwigii, Bacillus cereus, B. subtilis, Lactobacillus plantarum, and Saccharomyces cerevisiae). Twenty-four steers were blocked by initial BW ($634{\pm}16kg$) and randomly allocated to one of two treatments (control vs. 3.3% TMC). The effect of TMC supplementation on the growth performance was not significant. There was no incidence of urolithiasis in TMC-fed steers. However 3 out 12 steers (25%) fed the control diet were observed to have urinary calculi. The carcass yield and meat quality parameters were not affected by TMC supplementation, however marbling score was increased in TMC-fed steers (P = 0.08). There was no effect of TMC treatment on the chemical composition of longissimus dorsi muscle (LM). The TMC supplementation increased concentrations of manganese (P < 0.01), cobalt (P = 0.02), iron, and copper (P = 0.06) in LM. In conclusion, TMC treatment did not negatively affect growth performance and meat quality parameters, and positively affected the trace minerals profile of LM.

• Microbiology and Biotechnology Letters
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• v.29 no.1
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• pp.1-11
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• 2001

Lactic acid bacteria (LAB) are widely used for various food fermentation. With the recent advances in modern biotechnology, a variety of bio-products with the high economic values have been produced using microorganisms. For molecular cloning and expression studies on the gene of interest, E. coli has been widely used mainly because vector systems are fully developed. Most plasmid vectors currently used for E, coli carry antibiotic-resistant markers. As it is generally believed that the antibiotic resistance markers are potentially transferred to other bacteria, application of the plasmid vectors carrying antibiotic resistance genes as selection markers should be avoided, especially for human consump-tion. By contrast, as LAB have some desirable traits such that the they are GRAS(generally recognized as safe), able to secrete gene products out of cell, and their low protease activities, they are regarded as an ideal organism for the genetic manipulation, including cloning and expression of homologous and heterologous genes. However, the vec-tor systems established for LAB are stil insufficient to over-produce gene products, stably, limiting the use of these organisms for industrial applications. For a past decade, the two popular plasmid vectors, pAM$\beta$1 of Streptococcus faecalis and pGK12 theB. subtilis-E. coli shuttle vector derived from pWV01 of Lactococcus lactis ssp. cremoris wg 2, were most widely used to construct efficient chimeric vectors to be stably maintained in many industrial strains of LAB. Currently, non-antibiotic markers such as nisin resistance($Nis^{r}$ ) are explored for selecting recombi-nant clone. In addition, a gene encoding S-layer protein, slp/A, on bacterial cell wall was successfully recombined with the proper LAB vectors LAB vectors for excretion of the heterologous gene product from LAB Many food-grade host vec-tor systems were successfully developed, which allowed stable integration of multiple plasmid copies in the vec-mosome of LAB. More recently, an integration vector system based on the site-specific integration apparatus of temperate lactococcal bacteriophage, containing the integrase gene(int) and phage attachment site(attP), was pub-lished. In conclusion, when various vector system, which are maintain stably and expressed strongly in LAB, are developed, lost of such food products as enzymes, pharmaceuticals, bioactive food ingredients for human consump-tion would be produced at a full scale in LAB.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.10
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• pp.1464-1468
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• 2005

The aim of this study was to assess the effects of dietary complex probiotic (Lactobacillus acidophilus, $1.0{\times}10^7$ CFU/g; Saccharomyces cerevisae, $4.3{\times}10^6$ CFU/g; Bacillus subtilis $2.0{\times}10^6$ CFU/g) on growth performance, nutrients digestibility, blood characteristics and fecal noxious gas content in growing pigs. Ninety [(Duroc${\times}$Yorkshire)${\times}$Landrace] pigs with the average initial BW of 39.75${\pm}$1.97 kg were allocated into three treatments by a randomized complete block design. There were five pens per treatment with six pigs per pen. Dietary treatments include: 1) CON (basal diet); 2) CP1 (basal diet+complex probiotic 0.1%) and 3) CP2 (basal diet+complex probiotic 0.2%). During the entire experimental period of 6 weeks, results showed that addition of complex probiotic at the level of 0.2% to diet increased ADG significantly (p<0.05). Also, digestibilities of DM and N tended to increase, however, no significant differences were observed (p>0.05). Blood characteristics (IgG, Albumin, total protein, RBC, WBC and lymphocyte) of pigs were not affected (p>0.05) by complex probiotic supplementation. Fecal $NH_3$-N was decreased (11.8%) significantly by the addition of complex probiotic (p<0.05), but no effects were observed on fecal acetic acid, propionic acid and butyric acid concentrations (p>0.05). In conclusion, results in this experiment indicated that dietary complex probiotic supplementation had a positive effect on growing pigs performance and could decrease fecal $NH_3$-N concentration.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.3
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• pp.1299-1306
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• 2014

Background: Nano-biotechnology is recognized as offering revolutionary changes in various fields of medicine. Biologically synthesized silver nanoparticles have a wide range of applications. Materials and Methods: Silver nanoparticles (AgNPs) were biosynthesized with an aqueous extract of Pterocladiella (Pterocladia) capillacea, used as a reducing and stabilizing agent, and characterized using UV-VIS spectroscopy, Fourier Transform Infra red (FT-IR) spectroscopy, transmission electron microscopy (TEM) and energy dispersive analysis (EDX). The biosynthesized AgNPs were tested for cytotoxic activity in a human hepatocellular carcinoma ($HepG_2$) cell line cultured in Dulbecco's modified Eagle medium supplemented with 10% fetal bovine serum, 1% antibiotic and antimycotic solution and 2 mM glutamine. Bacterial susceptibility to AgNPs was assessed with Staphylococcus aureus, Bacillus subtilis [Gram+ve] and Pseudomonas aeruginosa and Escherichia coli [Gram-ve]. The agar well diffusion technique was adopted to evaluate the bactericidal activity of the biosynthesized AgNPs using Ampicillin and Gentamicin as gram+ve and gram-ve antibacterial standard drugs, respectively. Results: The biosynthesized AgNPs were $11.4{\pm}3.52$ nm in diameter. FT-IR analysis showed that carbonyl groups from the amino acid residues and proteins could assist in formation and stabilization of AgNPs. The AgNPs showed potent cytotoxic activity against the human hepatocellular carcinoma ($HepG_2$) cell line at higher concentrations. The results also showed that the biosynthesized AgNPs inhibited the entire panel of tested bacteria with a marked specificity towards Bacillus subtillus. Conclusions: Cytotoxic activity of the biosynthesized AgNPs may be due to the presence of alkaloids present in the algal extract. Our AgNPs appear more bactericidal against gram-positive bacteria (B. subtillus).

• Microbiology and Biotechnology Letters
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• v.30 no.3
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• pp.247-252
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• 2002

A bacterium producing the extracellular mannanase was isolated from Korean formented food and has been identified as a member of the genus Bacillus from the result of the phylogenic analysis based on partial 165 rRNA sequences. The isolate, named Bacillus sp. WL-3, was shown to be similar to B. subtilis strain on the basis of its biochemical properties. The mannanase of culture supematant was the most active at $55^{\circ}C$ and pH 6.0. The additional carbohydrates including u-cellulose, avicel, oat spelt xylan, guar gum and locust bean gum (LBG) increased the mannanase productivity. Especially, the maximum mannanase productivity was reached 65.5 U/ml in LB medium supplemented with 0.5% (w/v) LBG, which was 131-folds more than that in LB medium. It was sug-gested that the increase of mannanase production was owing to induction of mannanase biosynthesis by LBG hydrolysates transported following initial hydrolysis by extracellular mannanase during the cell growth. The molec-ular weight of WL-3 mannanase was estimated to approximately 38.0 kDa by zymogram on SDS-PAGE.

• Journal of Microbiology and Biotechnology
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• v.23 no.9
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• pp.1221-1228
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• 2013

Two lipase genes (bpl1 and bpl3) from Antarctic Bacillus pumilus strains were expressed in Bacillus subtilis. Both recombinant lipases BPL1 and BPL2 were secreted to the culture medium and their activities reached 3.5 U/ml and 5.0 U/ml, respectively. Their molecular masses apparent using SDS-PAGE were 23 kDa for BPL1 and 19 kDa for BPL3. Both lipases were purified to homogeneity using ammonium sulfate precipitation and HiTrap SP FF column and Superose 12 column chromatographies. The final specific activities were estimated to be 328 U/mg for BPL1 and 310 U/mg for BPL3. Both lipases displayed an optimum temperature of $35^{\circ}C$, similar to other mesophilic enzymes. However, they maintained as much as 70% and 80% of the maximum activities at $10^{\circ}C$. Accordingly, their calculated activation energy at a temperature range of $10-35^{\circ}C$ was 5.32 kcal/mol for BPL1 and 4.26 kcal/mol for BPL3, typical of cold-adapted enzymes. The optimum pH of BPL1 and BPL3 was 8.5 and 8.0, respectively, and they were quite stable at pH 7.0-11.0, showing their strong alkaline tolerance. Both lipases had a preference toward medium chain length ($C_6-C_{10}$) fatty acid substrates. These results indicate the potential for the two Antarctic B. pumilus lipases as catalysts in bioorganic synthesis, food, and detergent industries.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.1
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• pp.39-47
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• 2009

Gaegurin 4(GGN 4), an antimicrobial peptide isolated from a Korean frog, is five times more potent against Gram-positive than Gram-negative bacteria, but has little hemolytic activity. To understand the mechanism of such cell selectivity, we examined GGN4-induced $K^+$ efflux from target cells, and membrane conductances in planar lipid bilayers. The $K^+$ efflux from Gram-positive M. luteus(2.5 ${\mu}g/ml$) was faster and larger than that from Gram-negative E. coli(75 ${\mu}g/ml$), while that from RBC was negligible even at higher concentration(100 ${\mu}g/ml$). GGN4 induced larger conductances in the planar bilayers which were formed with lipids extracted from Gram-positive B. subtilis than in those from E. coli(p<0.01), however, the effects of GGN4 were not selective in the bilayers formed with lipids from E. coli and red blood cells. Addition of an acidic phospholipid, phosphatidylserine to planar bilayers increased the GGN4-induced membrane conductance(p<0.05), but addition of phosphatidylcholine or cholesterol reduced it(p<0.05). Transmission electron microscopy revealed that GGN4 induced pore-like damages in M. luteus and dis-layering damages on the outer wall of E. coli. Taken together, the present results indicate that the selectivity of GGN4 toward Gram-positive over Gram-negative bacteria is due to negative surface charges, and interaction of GGN4 with outer walls. The selectivity toward bacteria over RBC is due to the presence of phosphatidylcholine and cholesterol, and the trans-bilayer lipid asymmetry in RBC. The results suggest that design of selective antimicrobial peptides should be based on the composition and topology of membrane lipids in the target cells.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2003.10b
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• pp.48-49
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• 2003

This project was conducted to development several camellia tea mixed herb teas having any physiological effects. Leaves of tea tree contain many compounds, such as polysaccharides, volatile oils, vitamins, minerals, purines, alkaloids(eg. caffeine) and polyphenols(catechins and flavonoids). Although all three tea types(green, oolonr and black) have antibacterial and free radical capturing(antioxidizing) activities, the efficacy decreases substantially the darker the variety of tea is. This is due to lower contents of anti-oxidizing polyphenols remaining in the leaves. Unlike tea tree(Camellia sinensis), the biochemical features and effects of camellia(Camellia japonica) are not well known. Fresh mature leaf of sasanqua camellia(C. sasanqua), roasted young leaf tea(C. japonica) and fresh mature leaf and bark of camellia had high antibacterial activity against P. vulgaris and B. subtilis. In antifungal activity bioassay, young leaf roasted teas of camellia and sasanqua camellia had high activity against C. albicans and T. beigelil. Plant extracts from Camelia japonica had higher inhibitory activity against fungi than against bacteria. In cytotoxic effect against human acute myelogenous leukaemia cell extracts including fresh leaf(200$\mu\textrm{g}$/m1), bark(230$\mu\textrm{g}$/ml) and flower tea (320$\mu\textrm{g}$/m1)inhibited growth of AML cells.(중략)