• 한국가축번식학회지
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• 제27권3호
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• pp.259-268
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• 2003

모유에 존재하는 유지방구의 막을 구성하는 주된 당단백질인 하나인 lactadherin(과거에는 BA46로 일컬어짐)은 rotavirus에 의한 감염증상을 예방하는 것으로 보고되고 있다. 본 연구에서는 retrovirus vector system을 이용하여 Chinese Hamster Ovary (CHO) 세포에 tetracycline에 의해 발현이 제어되는 promoter 하의 lactadherin 유전자를 전이 시킨 후 lactadherin이 tetracycline에 의해 발현이 유도되는지의 여부를 실험하였다. 먼저 기초 실험으로 대장균의 LacZ 유전자를 이용하여 tetracycline에 의한 유도 여부를 시험하였다. RevTet-On과 RevTRE-LacZ retrovirus를 동시감염시킨 NIH3T3는 doxycycline (tetracycline 유도체)에 의해 투여량에 비례하여 반응정도가 증가하는 양상을 나타내었으며 최대의 반응은 doxycycline 농도가 1 $\mu\textrm{g}$/ml 이상에서부터 관찰되었다. 이 예비실험의 결과를 바탕으로 RevTet-On과 RevTRE-Ltd retrovirus vectyor를 이용하여 사람의 lactadherin 유전자의 유도적 발현을 검정하였는데 CHO 세포에서 lactadherin 유전자의 유도적 발현을 RT-PCR 기법을 이용하여 확인하였다. 표적세포 내에서 외부에서 도입된 유전자가 지속적으로 발현될 경우 심각한 생리적 부작용을 야기시킨다는 사실을 감안할 때 본 실험의 결과는 유전자 치료와 형질전환동물의 생산에 크게 도움이 될 것으로 예상된다.

• 한국응용과학기술학회지
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• 제30권3호
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• pp.400-410
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• 2013

This study was carried out to investigate the effects of feeding the broilers that are exposed to extreme heat stress by control of inverse lighting times with night restricted feeding of extreme heat diet(EHD1, 2: extreme heat diet) containing different amount of soy oil, molasses, amino acids and vitamin C on short chain fatty acid and blood lipid profile. 300 broiler chickens(Abaica strain) were randomized into four dietary treatment groups according to a randomized block design on the day they were hatched. The four dietary treatment groups were: T1(EHD 1, 10:00~19:00 Dark, 19:00~10:00 Light), T2(EHD 2, 10:00~19:00 Dark, 19:00~10:00 Light), T3(EHD 1, 09:00~18:00 Dark, 18:00~09:00 Light), T4(EHD 2, 09:00~18:00 Dark, 18:00~09:00 Light). The body weight gain of the broilers was highest in T2, and high in order T1, T4, T3(p<0.05). Weights of the lymphoid organ, thymus and bursa of Fabricius were high in T1, T2 as compared to T3, T4 but spleen was lower in T4 than T1, T2, T3(p<0.05). Blood triglyceride, total cholesterol and glucose were higher in T1, T2 than T3, T4(p<0.05). LDL-C was high in orderT4, T3, T2, T1 but HDL-C showed the opposite trend(p<0.05). Blood concentrations of IgG, IgG and IgM were higher in T1, T2 than inT3, T4, but the corticosterone concentration decreased significantly in them. In T1 and T2, Lactobacillus in the feces increased, but total aerobic bacteria, E.coli, coliform bacteria was decreased rather significantly, compared with those in T3 and T4(p<0.05). Concentrations of acetic acid, propionic acid and total SCFA in cecum were high in order T2, T1, T3, T4, but butyric acid, isobutyric acid, valeric acid, isovaleric acid were lower in T1, T2 than in T3, T4 (p<0.05).

• Journal of Microbiology and Biotechnology
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• 제23권11호
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• pp.1519-1528
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• 2013

Bacillus pumilus SG2, a halotolerant strain, expresses two major chitinases designated ChiS and ChiL that were induced by chitin and secreted into the supernatant. The present work aimed to obtain a mutant with higher chitinolytic activity through mutagenesis of Bacillus pumilus SG2 using a combination of UV irradiation and nitrous acid treatment. Following mutagenesis and screening on chitin agar and subsequent formation of halos, the mutated strains were examined for degradation of chitin under different conditions. A mutant designated AV2-9 was selected owing to its higher chitinase activity. To search for possible mutations in the whole operon including ChiS and ChiL, the entire chitinase operon, including the intergenic region, promoter, and two areas corresponding to the ChiS and ChiL ORF, was suquenced. Nucleotide sequence analysis of the complete chitinase operon from the SG2 and AV2-9 strains showed the presence of a mutation in the catalytic domain (GH18) of chitinase (ChiL). The results demonstrated that a single base change had occurred in the ChiL sequence in AV2-9. The wild-type chitinase, ChiL, and the mutant (designated ChiLm) were cloned, expressed, and purified in E. coli. Both enzymes showed similar profiles of activity at different ranges of pH, NaCl concentration, and temperature, but the mutant enzyme showed approximately 30% higher catalytic activity under all the conditions tested. The results obtained in this study showed that the thermal stability of chitinase increased in the mutant strain. Bioinformatics analysis was performed to predict changes in the stability of proteins caused by mutation.

• 한국식품저장유통학회지
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• 제18권2호
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• pp.219-225
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• 2011

본 연구에서는 울금의 품종별 생리활성을 확인하기 위하여 우리나라에서 재배되고 있는 울금, 강황 및 보라울금의 근경을 실험재료로 하여 total polyphenol 함량, 항균활성, 항균물질의 열안정성 및 pH안정성과 항산화활성을 측정하였다. 울금 품종별 total polyphenol 함량은 울금과 강황에서 보라울금에 비하여 높은 함량을 나타내었다. 울금의 추출 용매에 따른 항균활성은 ethanol 추출물이 15.4~16.3 mm로 가장 강한 항균활성을 나타내었으며 ethylacetate, ether, hexane 순으로 높은 활성을 나타내었다. 품종별로는 울금 ethanol 추출물의 항균활성이 다른 품종에 비하여 높게 나타났으나 E coli와 젖산균 2종, 효모 2종에서는 항균활성이 나타나지 않았다. 울금 ethanol 추출물은 열 및 pH변화에 안정함을 확인하였다. 추출물별 울금의 전자공여능은 ethanol 추출물에서 74.2%로 높은 항산화성을 보였고 ethyl acetate, ether, water, hexane 추출물 순으로 나타났다. 품종별 울금의 전자공여능을 ethanol 추출물에서 비교했을 때 울금 74.2%, 강황 43.98%, 보라울금 43.05%로 나타나 울금에서 강한 항산화성을 보였다. 대조구로 사용한 vitamin C와 BHA는 각각 95%와 96.1%의 높은 전자공여능을 나타내었다. 울금 품종별 항산화성은 울금이 강황과 보라울금에 비하여 월등히 높음을 확인하였다.

• 미생물학회지
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• 제42권1호
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• pp.1-6
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• 2006

Saccharomyces cerevisiae Dna2 helicase/endonuclease는 진핵세포 DNA 복제과정의 Okazaki fragment processing에서 RNA primer를 제거하는데 필수적인 역할을 한다. Genome-wide scale의 면역침전 실험결과, 기능이 알려져 있지 않은 단백질인 YHR122W가 Dna2 단백질과 상호작용한다고 예측되었다 (1). 본 연구에서는 이를 확인하기 위하여 YHR122W 유전자를 효모에서 과량발현시킨 결과, $dna2\Delta405N$ 돌연변이의 온도감수성 표현형이 억제되는 유전학적 상호작용을 관찰하였다. YHR122W 단백질이 Dna2 단백질과 직접적인 삼호작용을 하는지 확인하기 위하여 YHR122W를 대장균에서 재조합 단백질로 발현시키고 단백질을 정제하였다. Enzyme-linked immunosorbent assay를 통한 분석에서 YHR122W 단백질과 Dna2 단백질 사이의 상호작용을 확인하였다. 뿐만 아니라 YHR122W-Dna2 상호작용은 생리적 염도인 150 mM NaCl농도에서 가장 강한 결합을 보였다. 이러한 유전학적 상호작용과 물리적인 상호작용은 YHR122W가 생체내에서 Dna2의 기능과 밀접한 연관이 있을 가능성을 제시하고 있다.

• 약학회지
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• 제37권3호
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• pp.235-242
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• 1993

8-Fluorociprofloxacin(8-FCP) is an investigational quinolone derivative that is substituted with fluorine at the C-8 position of ciprofloxacin(CP). It was found that the in vitro activity of 8-FCP against Gram(+) bacteria was more potent that of CP, but the opposite against Gram(-) bacteria was true. However, 8-FCP showed better in vivo efficacy than CP against representative Gram(-) organisms, E. coli and K pneumoniae. In an attempt to seek for factors causing this discrepancy in the antibacterial activities, a comparative pharmacokinetic study of 8-FCP and CP was conducted in mice and rats treated either intravenously or orally at a single dose of 30 mg/kg. The pharmacokinetic parameters in mice were as follows; the mean peak serum concentrations(C$_{max}$) following i.v. and oral doses were 12.4 and 5.3 $\mu\textrm{g}$/ml for 8-FCP, and 9.5 and 2.5 $\mu\textrm{g}$/ml for CP, respectively. The terminal half-life(t$_{1/2\beta}$) was 72.9 min for 8-FCP, and 98.2 min for CP, and the oral bioavailability(F) was 89.9% for 8-FCP, and 50.5% for CP. In rats, the mean ($\pm$SD) $C_{max}$ after i.v. administration were 11.6$\pm$1.6 $\mu\textrm{g}$/ml for 8-FCP, and 10.2$\pm$1.3 $\mu\textrm{g}$/ml for CP, whereas oral administration produced $C_{max}$ of 5.9$\pm$1.8 $\mu\textrm{g}$/ml for 8-FCP and 1.1$\pm$0.9 $\mu\textrm{g}$/ml for CP, respectively. The t$_{1/2\beta}$ was 67.9$\pm$8.4 min for 8-FCP, and 76.4$\pm$7.2 min for CP. The F was 88.6$\pm$6.3% for 8-FCP, and 40.7$\pm$6.5% for CP. Marked differences were observed between the two quinolones in the $C_{max}$ and the area under the concentration-time curve obtained after oral administration in mice and rats. The extent of 8-FCP absorption in both mice and rats was approximately 2-fold higher than that of CP, suggesting that the fluorine atom attached to C-8 plays an important role in facilitating oral absorption from the gastrointestinal tract.

• 식물병연구
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• 제12권3호
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• pp.254-259
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• 2006

버섯폐배지로부터 잡균의 증식을 최소화하기 위해 버섯폐배지현탁액을 nutrient agar(NA) 배지에 직접 혼합하여 분주한 결과 배양기에 출현한 균체수가 희석평판법에 의한 것보다 훨씬 적었으며 형태적으로 Bacillus 속 세균이 분리될 확률이 높았다. 대장균(E. coli) $DH5{\alpha}$에 대하여 항균력을 보이는 1차 분리 균주를 신령버섯폐배지에서 12균주, 양송이폐배지에서 10균주, 느타리폐배지는 20균주 등 총 42균주를 분리하였다. 1차 선발한 균주를 최소 배지상에서 8종의 중요 식물병원균과 대치 우수한 길항력을 나타내는 17개의 균주를 2차 선발하였는데 이 들 중에서 배지상에서 생장이 우수하고 중요 식물병원균에 대하여 넓은 길항력을 가지는 B23과 B42균주를 최종 선발하였다. 이들은 각각 Brevibaciilus brevis와 Bacillus stearothermophilus로 동정되었다. 이들 길항균의 효과를 높이고 안전화를 위해서 유기농업에서 활용가능한 전분과 글리세린 및 계란노른자를 보조제로 선발하였다. 길항균과 이들 보조제를 혼합할 경우 대부분의 처리에서 세균의 밀도가 30일간 효과적으로 유지되거나 증가되었다. B23 단독 처리의 경우에는 상추균핵병의 발병율이 약 28% 정도였으나 전분(starch)과 글리세롤(glycerol) 및 계란노른자(Egg yolk)를 각각 0.2% 혼합 처리시는 발병율이 5% 이하로 나타나 길항세균 단독으로 처리하였을 경우보다 상추균핵병원의 방제 상승효과를 확인하였다.

• 대한치과보철학회지
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• 제37권6호
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• pp.741-755
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• 1999

As a material of metal-ceramic prosthesis, nickel as a form of Ni-Cr alloy has been used for many dental prostheses in many cases. However, several problems in use of the alloy have been revealed (ex : tissue stimulation, skin allergy, hypersensitivity, cytotoxicity and carcinogenecity). Little is known about nickel with respect to the relationship between Ni-prosthesis and gaining of Ni-resistance in oral microorganisms. The present study was undertaken to check wheather use of Ni-prosthesis leads to occurrence of Ni-resistant microorganisms. So this study may suggest the possible relationships between the oral microorganisms and nickel-resistance in oral environment. Bacteria were isolated from the gingival crevicular fluid on the pateints wearing Ni-Cr prosthesis. The isolated bacteria were tested for their Ni-resistance in nickel containing media at different concentration from 3mM to 110mM. E. coli HB101 was used as control. The Ni-resistant bacteria were isolated and biochemically identified. The Ni-resistant bacteria were tested several bio-chemical, molecular-biological tests. Performed tests were ; measuring the growth curve, antibiotic test, growth ability test in liquid media, isolation of the chromosome and plasmid, digestion of DNA by restriction enzyme, electrophoresis of chromosome and plasmid DNA, identification of Ni-resistant genes by the DNA hybridization. The results were as follows: 1) The bacteria isolated from gingival crevicular fluid on the patients wearing Ni-Cr alloy pros-thesis showed nickel-resistance. 2) The isolated microorganisms grew at nickel containing media of high concentrations (60mM-110mM). 3) Based on the biochemical tests, the isolated microorganisms were identified as Enterococcus faecalis(13 cases), Klebsiella pneumoniae(1 case) and Enterobacter gergeviae(1 case). 4) Enterococcus faecalis expressed not only nickel resistance but also the multi-drug resistance to several antibiotics ; chloramphenicol, kanamicin, streptomycin, lincomycin, clindamycin. However, all strain showed the sensitivity against the tetracycline. 5) DNA hybridization result suggest that there is no homology between the previousely known gene of nickel resistance in Klebsiella pneumoniae and chromosomal DNA of Enterococcus faecalis.

• BMB Reports
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• 제36권6호
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• pp.545-551
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• 2003

A cDNA of bovine brain glutamate dehydrogenase (GDH) was isolated from a cDNA library by recombinant PCR. The isolated cDNA has an open-reading frame of 1677 nucleotides, which codes for 559 amino acids. The expression of the recombinant bovine brain GDH enzyme was achieved in E. coli. BL21 (DE3) by using the pET-15b expression vector containing a T7 promoter. The recombinant GDH protein was also purified and characterized. The amino acid sequence was found 90% homologous to the human GDH. The molecular mass of the expressed GDH enzyme was estimated as 50 kDa by SDS-PAGE and Western blot using monoclonal antibodies against bovine brain GDH. The kinetic parameters of the expressed recombinant GDH enzymes were quite similar to those of the purified bovine brain GDH. The $K_m$ and $V_{max}$ values for $NAD^+$ were 0.1 mM and $1.08\;{\mu}mol/min/mg$, respectively. The catalytic activities of the recombinant GDH enzymes were inhibited by ATP in a concentration-dependent manner over the range of 10 - $100\;{\mu}M$, whereas, ADP increased the enzyme activity up to 2.3-fold. These results indicate that the recombinant-expressed bovine brain GDH that is produced has biochemical properties that are very similar to those of the purified GDH enzyme.

• Journal of Microbiology and Biotechnology
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• 제23권6호
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• pp.818-825
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• 2013

We isolated and functionally characterized the ${\alpha}$- and ${\beta}$-subunits (ApCpnA and ApCpnB) of a chaperonin from Aeropyrum pernix K1. The constructed vectors pET3d-ApCpnA and pET21a-ApCpnB were transformed into E. coli Rosetta (DE3), BL21 (DE3), or CodonPlus (DE3) cells. The expression of ApCpnA (60.7 kDa) and ApCpnB (61.2 kDa) was confirmed by SDS-PAGE analysis. Recombinant ApCpnA and ApCpnB were purified by heat-shock treatment and anion-exchange chromatography. ApCpnA and ApCpnB were able to hydrolyze not only ATP, but also CTP, GTP, and UTP, albeit with different efficacies. Purified ApCpnA and ApCpnB showed the highest ATPase, CTPase, UTPase, and GTPase activities at $80^{\circ}C$. Furthermore, the addition of ApCpnA and ApCpnB effectively protected citrate synthase (CS) and alcohol dehydrogenase (ADH) from thermal aggregation and inactivation at $43^{\circ}C$ and $50^{\circ}C$, respectively. In particular, the addition of ATP or CTP to ApCpnA and ApCpnB resulted in the most effective prevention of thermal aggregation and inactivation of CS and ADH. The ATPase activity of the two chaperonin subunits was dependent on the salt concentration. Among the ions we examined, potassium ions were the most effective at enhancing the ATP hydrolysis activity of ApCpnA and ApCpnB.