• 미생물학회지
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• 제46권4호
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• pp.359-365
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• 2010

조릿대 근권토양으로부터 분리한 방선균 50균주를 대상으로 고추 세균성 점무늬병원균(Xanthomonas axonopodis pv. vesicatoria)의 항균활성 12균주를 선발하였다. 이들 항균활성 12균주의 계통학적 위치를 검토한 결과, 모두 Streptomyces 속의 Cluster II에 속하는 특징을 나타내었다. JR-24 균주는 최소저해 농도(MIC) 10 ${\mu}l$/disc를 나타내었으며, 배양액 5 ${\mu}l$/ml를 처리 하여 12시간 배양한 결과 Xanthomonas axonopodis pv. vesicatoria에 강한 생육저해효과를 나타내어 최우수 균주로 선발되었다. 항균활성 균주 JR-24의 16S rRNA 유전자 염기서열을 검토한 결과, Streptomyces galbus $DSM40089^T$ (X79852)와 98.1%, Streptomyces longwoodensis $LMG20096^T$ (AJ781356)와 98% 그리고 Streptomyces capoamus $JCM4734^T$ (AB045877)와 97.8%의 상동성을 나타내었다. API 20NE와 API 50CHE를 이용하여 JR-24 균주의 생리 생화학적 특성을 확인한 결과, L-arabinose, D-fructose, D-glucose, D-galactose을 이용하며 gelatin, protein, starch에 대하여 분해능이 있는 것으로 확인되었다. 주요지방산으로는 iso-$C_{14:0}$ (25.93%), iso-$C_{15:0}$ (10.13%), anteiso-$C_{15:0}$ (19.29%) 그리고 iso-$C_{16:0}$ (20.35%) 등을 함유하였으며, 퀴논종은 MK-9 ($H_4$) 4.37%, MK-9 ($H_6$) 51.22% 그리고 MK-9 ($H_8$) 49.47%로 동정되었다. Streptomyces sp. JR-24 균주의 계통학적 특성을 근연종인 Streptomyces galbus $DSM40089^T$와 비교한 결과, 다수의 표현형적 및 계통학적 차이를 나타내었다. 본 연구에서 분리된 Streptomyces sp. JR-24는 친환경 미생물제제 개발을 위한 유전자원 확보에 있어서 매우 큰의의가 있을 것으로 사료 된다.

• Asian-Australasian Journal of Animal Sciences
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• 제17권6호
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• pp.863-868
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• 2004

Lectin activities and chemical characteristics of Escherichia coli, Lactobacillus spp. and Bifidobacterium spp. originating from the porcine cecal mucosal layer were studied based on hemagglutination assay (HA) and hemagglutination inhibition assay (HIA). Although all the bacterial strains were able to agglutinate erythrocytes of porcine or rabbit origin, much higher HA titers were consistently observed for Lactobacillus spp. than for E. coli or for Bifidobacterium spp. A remarkable reduction in HA titers occurred by the treatment of E. coli and Lactobacillus spp. with protease or trypsin and of Bifidobacterium spp. with protease, trypsin or periodate. There were no significant effects on the HA titers of the three groups of bacteria after the treatment with lipase. Hemagglutination of E. coli was strongly inhibited by D (+)-mannose and D (+)-galactose; Lactobacillus spp. by $\alpha$-L-rhamnose and methyl-$\beta$-galactopyranoside; Bifidobacterium spp. by D (+)-alactose, $\alpha$-L-rhamnose, $\alpha$-L-fucose, L (+)-arabinose, D (+)-mannose, D (-)-fructose at a relatively low concentration (1.43 to 3.75 mg/ml). These results, combined with the enhanced HA activities of the three bacterial strains by modification of rabbit erythrocytes with neuraminidase and abolished HA activity of E. coli after treatment with $\beta$-galactosidase, indicate that it might be the glycoproteinous substances surrounding the surface of the bacterial cells that are responsible for the adhesions of these microorganisms by recognizing the specific receptors on the red blood cell.

• Journal of Applied Biological Chemistry
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• 제48권1호
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• pp.7-10
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• 2005

Trichoderma sp. SY most effectively produces an extracellular ${\gamma}$-L-arabinofuranosidase (AF) using arabinose as a carbon source. AF grown on cellulose as a carbon source was purified 28-fold with 4.4% yield by DEAE exchange and HQ/20 cation exchange chromatographies The purified enzyme was found to be homogeneous on SDS-PAGE with molecular weight of 89 kDa. It exhibited a high level of activity with p-nitrophenyl ${\alpha}$-L-arabinofuranoside, showing $K_m$ and $V_{max}$ values of $0.15\;{\mu}M$ and $239.85U{\cdot}mg^{-1}$, respectively and did not require any metal ion for activity. It also released p-nitrophenol from p-nitrophenol conjugated ${\beta}$-D-xylopyranoside, and ${\beta}$-D-galactopyranoside not from ${\beta}$-D-glucopyranoside.

• 한국미생물·생명공학회지
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• 제22권6호
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• pp.607-613
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• 1994

The Bacillus stearothermophilus arfI gene encoding a-arabinofuranosidase was isolated from the genomic library, cloned into pBR322, and subsequently transferred into the Escherichia coli HB101. The recombinant E. coli was selected from approximately 10,000 transformants screened by making use of its ability to produce a yellow pigment around the colony on the selective medium supplemented with p-nitrophenyl-$\alpha$-L-arabinofuranoside (pNPAf), a chromogenic substrate. The functional clone was found to harbor a recombinant plasmid, pKMG11 with an insertion of about 5 kb derived from the B. stearothermophilus chromosomal DNA. Identity of the arfI gene on the insert DNA was confirmed by a zymogram with 4-methylumbelliferyl-$\alpha$-L-arabinofuranoside as the enzyme substrate. The $\alpha$-arabinofuranosidase from the recombinant E. coli strain showed very high substrate specificity; the enzyme displayed high activity only with pNPAf among many other p- or $o$-nitrophenyl derivatives of several sugars, and acted only on arabinoxylan among various natural arabinose containing polysaccharides tested.

• 한국식품과학회지
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• 제34권5호
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• pp.879-884
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• 2002

감잎으로부터 항보체 활성물질을 분리정제하기 위해 감잎 (250 g)을 $100^{\circ}C$에서 3시간 동안 열수 추출하고 분자량 10 kDa membrane을 사용하여 농축한 후 ethanol 침전과 methanol 추출을 통해 조다당류(DKC)를 얻었다. 조다당류의 정제는 DEAE-Toyopearl 650C와 Bio-gel P60을 사용하여 실시하였다. 정제된 DKC-1c는 $1000\;{\mu}g/mL$의 농도에서 고전경로를 통해서는 85.4% 활성화시켰고 부경로에서는 65.1% 활성화시켰다. 정제 다당류 DKC-1c는 분자량은 66.6 kDa이고 정제도가 높은 중성다당류로써 주요 구성당은 glucose(29.0 mol.%), arabinose(24.3 mol.%), galactose(16.2 mol.%) 순으로 검출되었다. 면역전기영동을 통하여 확인한 결과 DKC-1c는 C3를 부경로에서도 C3a와 C3b로 활성화시키는 complement activator임이 확인되었다.

• Journal of Microbiology and Biotechnology
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• 제17권3호
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• pp.481-489
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• 2007

[ $\alpha$ ]-L-Arabinofuranosidases (AFases; EC 3.2.1.55) are exo-type enzymes, which hydrolyze terminal nonreducing arabinose residues from various polysaccharides such as arabinan and arabinoxylan. Genome-wide BLAST search showed that various bacterial strains possess the putative AFase genes with well-conserved motif sequences at the nucleotide and amino acid sequence levels. In this study, two sets of degenerate PCR primers were designed and tested to detect putative AFase genes, based on their three highly conserved amino acid blocks (PGGNFV, GNEMDG; and DEWNVW). Among 20 Bacillus-associated species, 13 species were revealed to have putative AFase genes in their genome and they share over 67% of amino acid identities with each other. Based on the partial sequence obtained from an isolate, an AFase from Geobacillus sp. was cloned and expressed in E. coli. Enzymatic characterization has verified that the resulting enzyme corresponds to a typical AFase. Accordingly, degenerate PCR primers developed in this work can be used for fast, easy, and specific detection and isolation of putative AFase genes from bacterial cells.

• Food Science and Biotechnology
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• 제17권5호
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• pp.990-995
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• 2008

A thermostable $\alpha$-L-arabinofuranosidases ($\alpha$-L-AFase) is an industrially important enzyme for recovery of L-arabinose from hemicellulose. The recombinant $\alpha$-L-AFase from Thermotoga maritima was expressed in Escherichia coli by using a constitutive pHCE or an inducible pRSET vectors. In batch fermentation, the constitutive expression system resulted in slightly faster growth rate (0.78 vs. 0.74/hr) but lower enzyme activity (2,553 vs. 3,723 units/L) than those of the induction system. When fed-batch fermentation was performed, biomass and enzyme activity reached the highest levels of 36 g/L and 9,152 units/L, respectively. The fed batch cultures performed superior results than batch culture in terms of biomass yield (4.62-5.42 folds) and enzyme synthesis (3.39-4.00 folds). In addition, the fed-batch induction strategy at high cell density resulted in the best productivity in cell growth as well as enzyme activity rather than the induction method at low cell density or the constitutive expression.

• 한국식품영양학회지
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• 제30권6호
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• pp.1140-1148
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• 2017

For the purpose of developing new immunomodulatory agents from broccoli, ethanol extract (BCEE), hot water extract (BCHW), and crude polysaccharide (BCCP) were isolated from broccoli, and their immunomodulatory activities and chemical properties were examined. In the in vitro cytotoxicity analysis, BCHW and BCCP did not affect the growth of tumor cells and normal cells. Murine peritoneal macrophages stimulated with BCCP showed higher production of IL-6, IL-12, and $TNF-{\alpha}$ cytokines than those stimulated with BCHW. Also, BCHW and BCCP did not show proliferation of splenic lymphocytes. In the in vitro assay for intestinal immunomodulatory activities, only BCCP enhanced GM-CSF secretion and the bone marrow cell-proliferating activity via cells in Peyer's patches at $1,000{\mu}g/mL$. Also, BCHW mainly contained 33.7% neutral sugars, such as arabinose, glucose, and galactose, and 30.7% uronic acid, and BCCP consisted of 42.6% neutral sugars, including arabinose, galactose, and glucose, and 50.5% uronic acid. The above results lead us to conclude that crude polysaccharide (BCCP) isolated from broccoli causes considerably high cytokine production in peritoneal macrophages and bone marrow cell proliferation, and the polysaccharide extraction process is indispensable for separation of new immunomodulatory agents from broccoli.

• Applied Biological Chemistry
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• 제39권2호
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• pp.159-164
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• 1996

혈액 항응고 활성물질을 검색하기 위하여 73종의 한약재, 41종의 산채류 및 일반 채소류, 5종의 해조류들의 열수추출물들에 대해 혈액응고 지연시간(Tr)을 시험하였다 1차검색에서 비교적 활성이 높았던 택사, 삼칠근, 현호색, 운지, 마늘, 청각의 산성 수추출물에서 가장 높은 항응고 활성(대조군에 비해 4.3배)을 보였다. 청각(1.2KG)을 0.8% 염산 수용액(24l)으로 추출한 후 메탄을 환류, 에탄올 환류, 침전 및 투석하여 활성이 약 2배 증가된 조다당 CF-1을 얻었다. CF-1은 80.8%의 다당과, 8.7%의 단백질 및 13.3%의 황산기로 구성되어 있었으며, 다당류는 arabinose, galactose, 및 glucose가 주구성당으로 확인되었다. 이 조다당 CF-1의 항응고 활성 본체를 확인하기 위해 pronase처리와 periodate 산화를 행한 결과 pronase 처리시에는 활성의 변화가 없었으나 periodate 산화시에는 약 70%로 활성이 감소한다는 사실로부터 CF-1의 항응고 활성은 다당에 의해 기인됨을 알 수 있었다.

• 한국미생물·생명공학회지
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• 제35권3호
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• pp.191-195
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• 2007

탄닌 분해균 선발배지를 이용하여 탄닌 분해활성 보유 유산균을 발효 침채류로부터 분리하는 과정에서 오이소박이, 파김치, 총각김치로부터 다당류로 추정되는 점질물질을 생산하는 균주 4종을 선발하였다. 이들 균주의 165 rDNA 염기서열에 의한 계통유전학적 동정 결과, CK31은 Enterobacter cowanii, CK32는 Escherichia senegalensis, YK5는 Enterobacter asburiae, GK23은 Enterobacter ludwigii와 99% 이상의 매우 높은 상동성을 나타내었다. 가장 많은 양의 점질 물질을 세포 외로 생성하는 CK3l 균주는 rhamnose, fucose, arabinose, mannose, galactose, glucose의 중성당 조성을 가지고 있는 분자량 약 420 kDa heteropolysaccharide를 생산하는 것으로 나타났다.