• Journal of Microbiology and Biotechnology
• /
• 제28권11호
• /
• pp.1883-1895
• /
• 2018

Alcohol dependence is a global public health problem, yet the mechanisms of alcohol dependence are incompletely understood. The traditional view has been that ethanol alters various neurotransmitters and their receptors in the brain and causes the addiction. However, an increasing amount of experimental evidence suggests that gut microbiota also influence brain functions via gut-to-brain interactions, and may therefore induce the development of alcohol use disorders. In this study, a rat model of alcohol dependence and withdrawal was employed, the gut microbiota composition was analyzed by high-throughput 16S rRNA gene sequencing, and the metagenome function was predicted by PICRUSt software. The results suggested that chronic alcohol consumption did not significantly alter the diversity and richness of gut microbiota in the jejunum and colon, but rather markedly changed the microbiota composition structure in the colon. The phyla Bacteroidetes and eight genera including Bacteroidales S24-7, Ruminococcaceae, Parabacteroides, Butyricimonas, et al were drastically increased, however the genus Lactobacillus and gauvreauii in the colon were significantly decreased in the alcohol dependence group compared with the withdrawal and control groups. The microbial functional prediction analysis revealed that the proportions of amino acid metabolism, polyketide sugar unit biosynthesis and peroxisome were significantly increased in the AD group. This study demonstrated that chronic alcohol consumption has a dramatic effect on the microbiota composition structure in the colon but few effects on the jejunum. Inducement of colonic microbiota dysbiosis due to alcohol abuse seems to be a factor of alcohol dependence, which suggests that modulating colonic microbiota composition might be a potentially new target for treating alcohol addiction.

• 한국미생물·생명공학회지
• /
• 제30권4호
• /
• pp.312-319
• /
• 2002

저온성 균인 Sporosarcina psychrophilia의 염색체 DNA를 추출하여 Sau3AI으로 부분 절단하고 pUC19 vector에 ligation 시킨 후, Escherichia coli pyrB mutant 균주에 형질전환하여 uracil이 없는 AB배지에서 생존하는 균주를 선택한 후, 그 plasmid를 분리하여 pSMI과 pSM2라고 명명하였다. 두 plasmid의 염기배열을 결정한 결과 pSM2 insert DNA는 pSMl insert DNA 부분을 포함하는 2,606 nucleotide 단편이었다. 염기서열을 분석하였을 때 이것은 1개의 완전한 open reading frame(ORF)과 2개의 부분 ORFs를 포함하고 있었다. 두 번째 위치한 완전한 ORF는 Bacillus caldolyticus aspartate transcarbamylase(pyrB)와 아미노산 서열 수준에서 59% 상동성을 보였고, 첫 번째와 세 번째 위치한 부분적 ORFs는 각각 Bacillus속의 uracil permease(pyrP)와 dihydoorotase(pyrC)와 높은 상동성을 보였다. 그리고 pyrB와 pyrP사이에 intergenic 부분에는 잠재적인 terminator, antiterminator, anti-antiterminator 구조를 포함하고 있었다. 이러한 결과는 S. psychrophilia pyrimidine 생합성에 관련된 유전자들은 다른 Bacillus속에서 알려진 바와 같이 유전자군을 형성하고 있을 것으로 추정했다. S. psychrophilia pyrB 유전자의 생성물을 과다발현 시키고 정제해서 그 단백질을 SDS-PAGE로 확인한 결과 27 kDa 부근에서 band를 확인할 수 있었으며, 정제한 단백질도 ATCase 효소활성을 지니고 있었다.

• Journal of Microbiology and Biotechnology
• /
• 제12권1호
• /
• pp.114-120
• /
• 2002

A 1,989-bp genomic region encoding nickel resistance genes was isolated from Legionella pneumophila, a pathogen for legionellosis. From a sequencing and computer analysis, the region was found to harbor two structural genes, a nreB-like protein gene (1,149 bp) and a nreA-like protein gene (270 bp), in a row. Both genes exhibited a significant degree of similarity to the corresponding genes from Synechocystis sp. PCC6803 ($54\%$ amino acid sequence identity) and Achromobacter xylosoxidans 31A ($76\%$). The gene was successfully expressed in E. coli MG1655 and conferred a nickel resistance of up to 5 mM in an LB medium and 3 mM in a TMS medium including gluconate as the sole carbon source. E. coli harboring the nickel resistance gene also exhibited a substantial resistance to cobalt, yet no resistance to cadmium or zinc. Since the extracellular concentration of nickel remained constant during the whole period of cultivation, it was confirmed that the nickel resistance was provided by an efflux system like the $Ni^2＋$permease (nrsD) of Synechocystis sp. strain PCC6803. Since polyphosphate (poly-P) is known as a global regulator for gene expression as well as a potential virulence factor in E. coli, the nickel resistance of a ppk mutant of E. coli MG 1655 harboring the nickel resistance gene from L. pneumophila was compared with that of its parental strain. The nickel resistance was significantly attenuated by ppk inactivation, which was more pronounced in an LB medium than in a TMS medium.

• 미생물학회지
• /
• 제48권2호
• /
• pp.79-86
• /
• 2012

ErmSF는 23S rRNA의 A2058 (E. coli numbering)에 methylation을 유발하여 macrolide-lincosamide-streptogramin B ($MLS_B$)계 항생제의 부착을 저해함으로써 항생제 활성을 억제하는 내성인자 단백질인 Erm 단백질들 중의 하나이다. Erm 단백질들 사이에서 공통적으로 나타나는 $^{222}FXPXPXVXS^{230}$ (ErmSF numbering) 서열은 Erm 단백질인 ErmC'와 DNA methyltransferase인 M. Taq I의 구조를 분석한 연구에서 타깃인 adenine과 직접적으로 상호작용하는 부위로 제안되거나 확인되었다. 따라서 이 부분 중 Erm 단백질 사이에서 잘 보존되어있지는 않지만 염기성인 잔기의 특성상 기질인 RNA와 상호작용이 예상되는 223, 227번 arginine을 alanine으로 위치 지정 치환한 변이 단백질을 이용하여 그 잔기의 효소 활성에서의 역할을 확인하였다. 두 변이 단백질은 생체 내에서 그 활성을 여전히 유지하고 있어서 항생제인 erythromycin에 대하여 내성을 나타내었으나 in vitro 상에서는 R223A 또는 R227A가 야생형 ErmSF에 비하여 약 50%, 88%의 활성을 각각 나타내어 효소 활성에서 각 잔기가 결정적이지는 않지만 중요한 역할을 수행하고 있음을 확인하였다.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
• /
• 제26권6호
• /
• pp.581-590
• /
• 2000

Alterations in the cellular genome affecting the expression or function of genes controlling cell growth and differentiation are considered to be the main cause of cancer. Over 30 oncogenes can be activated by insertional mutagenesis, single point mutations, chromosomal translocations and gene amplification. The ras oncogenes have been detected in $15{\sim}20%$ of human tumors that include some of the most common forms of human neoplasia and are known to acquire their transforming properties by single point mutations in two domains of their coding sequences, most commonly in codons 12 and 61. The ras gene family consists of three functional genes, N-ras, K-ras and H-ras which encode highly similar proteins of 188 or 189 amino acid residues generically known as P21. ras proteins have been shown to bind GTP and GTP, and possess intrinsic GTPase activity. Experimental study was performed to observe the mutational change of the ras gene family and apply the results to the clinical activity. 36 Golden Syrian Hamster each weighing $60{\sim}80g$ were used and painted with 0.5% DMBA by 3 times weekly on the right buccal cheek(experimental side) for 6, 8, 10, 12, 14 and 16 weeks. Left buccal cheek (control side) was treated with mineral oil as the same manner of the right side. The hamsters were sacrificed on the 6, 8, 10, 12, 14 & 16 weeks. Normal and tumor tissues from paraffin block were completely dissected by microdissection and DNA from both tissue were isolated by proteinase K/phenol/chloroform extraction. Segments of the K-ras and H-ras gene were amplified by PCR using the oligonucleotide primers corresponding to the homologous region (codon 12 and 61) of the hamster gene, and then confirmational change of ras genes was observed by SSCP and autosequencing analysis. The results were as follows : 1. Malignant lesion could be found in the experimental side from the experimental six weeks. 2. One hamster among six showed point mutation of the H-ras codon 12($G{\rightarrow}A$ transition) at the experimental 10 and 14 weeks. 3. One of six at 6 weeks, two of six at 8 weeks and one of six at 12 weeks revealed the confirmational change of the H-ras codon 61($A{\rightarrow}T$ transversion). 4. The incidence of point mutation of H-ras codon 12 and 61 were 5.5%(2 of 36) and 11%(4 of 36) respectively. 5. Point mutation of the K-ras could not be seen during the whole experimental period. Form the above results, these findings strongly support the concept that H-ras oncogenes may have the influence of the DMBA induced carcinoma of hamster buccal pouch.

• 한국환경과학회지
• /
• 제25권4호
• /
• pp.579-588
• /
• 2016

본 연구에서는 홍어껍질에서 추출한 콜라겐 펩타이드 이화학적 특성과 비만 유발 쥐에 대한 투여 실험을 통해서 지방조직의 지질 감소효과를 조사하였으며, 홍어 껍질 콜라겐 펩타이드를 원료로 한 응용 제품 개발을 통하여 산업적 적용을 검토하였다. 일반성분 분석 결과 홍어 껍질 콜라겐 펩타이드는 대부분이 단백질로 이루어져 있는 고단백 저지방 물질이며, 분자량 측정 결과 약 1000 Da.의 저분자량이고, 아미노산 분석에서는 전형적인 콜라겐 단백질의 특성을 나타내었다. 비만 유발 쥐에 홍어 껍질 콜라겐 펩타이드를 500 mg/kg씩 18일간 경구 투여한 실험 결과에서 사료 섭취량과 음수량이 상당량 줄어 들었으며, 지방조직의 중성지방과 총 콜레스테롤 수치가 유의적으로 감소하였다. 홍어 껍질 콜라겐 펩타이드를 대량생산을 한 결과 제품의 최종 수율은 원료대비 약 17.23%로 상당히 높은 수율을 나타내었다. 이를 원료로 한 기능성 화장품 및 생활용품을 생산 및 출시하였다. 따라서 홍어 가공 부산물인 껍질을 재이용한 기능성 단백질 소재 개발을 통하여 건강기능식품과 화장품 분야에서의 기능성과 가격경쟁력을 갖춘 소재 개발에 기여 할 것으로 기대된다.

• 한국양식학회지
• /
• 제17권1호
• /
• pp.1-7
• /
• 2004

A rainbow trout uncoupling protein 3 (UCP3) cDNA clone, encoding a 310 amino acid protein, was cloned and sequenced from a liver cDNA library. Two different splice variants designated UCP3-vl and UCP3-v2, were identified through liver cDNA library screening using rainbow trout UCP3 cDNA clone as a probe. UCP3-vl has 3 insertions in the UCP3 cDNA: the first insertion (133 bp), the second (141 bp), and the third (370 bp) were located 126 bp, 334 bp and 532 bp downstream from the start codon, respectively. UCP3-v2 contained a single insertion, identical in sequence and location to the second insertion of UCP3-vl. UCP3, a mitochondrial protein, functions to modulate the efficiency of oxidative phosphorylation. UCP3 has been detected from heart, testis, spinal cord, eye, retina, colon, muscle, brown adipose tissue and white adipose tissue in mammalian animals. Human and rodent UCP3s are highly expressed in skeletal muscle and brown adipose tissue, while they show weak expression of UCP3 in heart and white adipose tissue. In contrast to mammalian studies, RT-PCR and Southern blot analysis of the rainbow trout demonstrated that UCP3 is strongly expressed in liver and heart. UCP3, UCP3-vl, and UCP3-v2 all contain an Ava III short interspersed element (SINE), located in the 3'untraslated region (UTR). PCR using primers from the Ava III SINE and the UCP3 3'UTR region indicates that the UCP3 cDNA is structurally conserved among salmonids and that these primers may be useful for salmonid species genotyping.

• 한국임상약학회지
• /
• 제20권3호
• /
• pp.235-241
• /
• 2010

Bambuterol hydrochloride, dimethylcarbamic acid 5-[2-(1,1-dimethylethyl)amino-1-hydroxyethyl]-1,3-phenylene ester hydrochloride, is the prodrug of active ${\beta}_2$-adrenergic metabolite terbutaline. The purpose of the present study was to evaluate the bioequivalence of two bambuterol hydrochloride tablets, $Bambec^{(R)}$ tablet 10 mg (Yuhan Co., Ltd.) and Bambucol tablet 10 mg (Sam Chun Dang Pharm. Co., Ltd.), according to the guidelines of Korea Food and Drug Administration (KFDA). In vitro release of bambuterol from two bambuterol hydrochloride formulations was tested using KP VIII Apparatus II method with various dissolution media. Twenty eight healthy male Korean volunteers, $23.86{\pm}1.65$ years in age and $68.98{\pm}9.58$ kg in body weight, were divided into two groups and a randomized $2{\times}2$ cross-over study was employed. After two tablets containing 10 mg as bambuterol hydrochloride were orally administered, blood samples were taken at predetermined time intervals, and the concentrations of bambuterol in serum were determined using column switching HPLC with UV detector. The dissolution profiles of two formulations were similar in all tested dissolution media. The pharmacokinetic parameters such as $AUC_t$, $C_{max}$ and $T_{max}$ were calculated, and ANOVA test with K-BE Test 2002 was utilized for the statistical analysis of the parameters using logarithmically transformed $AUC_t$, $C_{max}$ and untransformed $T_{max}$. The results showed that the differences between two formulations based on the reference drug, $Bambec^{(R)}$, were -8.10%, -3.82% and 12.65% for $AUC_t$, $C_{max}$ and $T_{max}$, respectively. There were no sequence effects between two formulations in these parameters. The 90% confidence intervals using logarithmically transformed data were within the acceptance range of log 0.8 to log 1.25 (i.e., log 0.8093~log 1.0302 and log 0.8564~log 1.1280 for $AUC_t$ and $C_{max}$, respectively). Thus, the criteria of the KFDA bioequivalence guideline were satisfied, indicating Bambucol tablet 10 mg was bioequivalent to $Bambec^{(R)}$ tablet 10 mg.

• 한국산림과학회지
• /
• 제102권3호
• /
• pp.331-337
• /
• 2013

Formate dehydrogenase(FDH)는 포름산이온을 이산화탄소로 산화하는 반응을 촉매하는 효소로서, 건조와 저온 그리고 병원균 감염 등에 반응하는 스트레스 단백질로 알려져 있다. 본 연구에서는 현사시나무에서 FDH의 cDNA를 분리하여 구조와 발현 특성 등을 조사하였다. 현사시나무의 FDH cDNA(PagFDH1)는 1,499개의 염기쌍으로 이루어져 있으며, 388개의 아미노산으로 구성되는 예상 분자량 42.5 kDa의 단백질을 암호화한다. PagFDH1 단백질은 미토콘드리아 신호펩티드와 $NAD^+$ 결합부위를 가지고 있다. PagFDH1은 현사시나무의 염색체에 1 copy가 존재하며, 배양세포에서 가장 높게 발현되고 뿌리와 꽃 그리고 잎에서도 발현되었다. 현탁배양세포의 생장주기에서 유도기와 초기 지수생장기에 높게 발현하였다. PagFDH1은 건조와 염 스트레스에 반응하여 ABA를 경유한 신호전달경로에 의해 발현이 유도되는 것으로 나타났다. 본 연구결과는 FDH 유전자의 도입과 발현조절을 통한 환경 스트레스 저항성나무의 개발에 도움을 줄 것으로 생각된다.

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
• /
• pp.749-754
• /
• 2000

알려진 dTDP-D-glucose 4,6-dehydratase의 amino acid 서열로 부터 primer를 제작하여 내열성 균주인 Thermus caldophilus GK24에서 colony hybridization 과정을 거쳐 dTDP-D-glucose 4,6-dehydratase를 포함하는 cosmid DNA를 얻었다. 유전자 분석을 위해 cosmid DNA를 subclone 하여 작은 크기로 분리하였다. 분리된 cosmid를 pSMTC-1 으로 명명하고 pSMTC-1를 BamHI으로 반응시켜 BamHI 단편 모두를 pGEM 7(+)를 이용하여 subclone 하였다. 각각의 이름은 크기에 따라 pKCB10(1.2kb-BamHI), pKCB20(1.6kb-BamHI), pKCB30(2.Ikb-BamHI), pKCB40(2.5kb-BamHI), pKCB50(2.5kb-BamHI), pKCB60(2.7kb-BamHI), pKCB70(3.4kb-BamHI), pKCB80(4.4kb-BamHI), pKCB90(7.0kb-BomHI) 으로 명명하였다. 각각의 subclone된 유전자를 분석하기 위해 Erase-a-base 방법을 이용하여 template를 준비하였고 이를 자동 염기서열 분석기를 이용하여 염기서열을 분석하였다. 염기서열분석 결과 pKCB80(4.2kb)에 dTDP-D-glucose synthase(orfA) 유전자를 비롯하여 dTDP-D-glucose 4,6-dehydratase(orfB), orfC (dTDP-4-keto-L-rhamnose reductase) 그리고 orfD(dTDP-4-keto-6-deoxy-D-glucose 3,5-epimerase)와 유사한 유전자들이 있음이 확인 되었고 dTDP-L-rhamnose의 생합성 과정을 예상할 수 있었다.