• 미생물학회지
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• 제37권3호
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• pp.221-227
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• 2001

열충격 단백질(heat shock protein: HSP)은 변성된 단백질의 응집을 방지하여 가혹한환경에서 병원균의 생존을 증가시킨다. 세균에 알코을 stress를 가하면 다량의 DnaK와 GronEL이 유도되지만 폐렴구균에서는 DnaK와 GroEL이 전혀 유도되지 않는 대신 알코올탈수소효소(alcohol dehydrogenase : ADH)가 유도되었다. 이런 특성은 폐렴구균 ADH가 HSP처럼 chaperone 기능을 수행라고 있을 가능성을 제시하고 있으므로 본 연구에서는 일차적으로 ADH 유전자를 확인하고 ADH 의 면역특성 및 세포내 분포를 측정하였다. 폐렴구균 ADH는 이질아메바 ADH2 및 대장균 ADH 와 높은 유사성을 나타냈으며 883 개의 아미노산으로 구성된 등전점 6.09의 단백질로 추정된다. 그러나 폐렴구균 ADH와 유사성이 높은 대장균, 유산균 및 황색포도상구균의 용해액을 폐렴구균 ADH 항체와 immunoblot을 실시하였을 때 전혀 반응하지 않았다. 또한 세포질, membrane, periplasm에 있는 단백질 분획 및 폐렴구균 배양 상등액을 ADH 항체와 immune blot을 실시하였을 때 ADH 는 열충격에 관계없이 세포 밖으로 분비되는 단백질임을 확인하였다. 이런 결과는 폐렴구균 ADH가 진단용항원 및 백신으로 개발될 수 있는 가능성을 제시하고 있다.

• 한국산학기술학회논문지
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• 제9권1호
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• pp.189-194
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• 2008

인진쑥 배지에서 배양한 노루궁뎅이버섯(HEAC), 노루궁뎅이 버섯과 인진쑥 추출물들의 음주 후 체내 알코올 농도의 감소 효과를 확인하였으며, 또한 혈중 알코올 탈수소효소 및 아세트알데히드 탈수소효소의 활성을 조사하여 알코올 분해 활성을 조사하였다. 체내 알코올 농도 측정결과 HEAC의 경우 170분, 노루궁뎅이 버섯 추출물은 210분후에 혈중 알코올이 검출되지 않았다. 또한, HEAC는 알코올 탈수소효소 활성에서 대조군 보다 시간에 따라 154%, 노루궁뎅이버섯 추출물은 148%로 비교적 높은 활성을 나타내었다. 아세트알데히드 탈수소활성에서는 HEAC와 노루궁뎅이버섯 추출물이 대조구에 비해 104-111%이상의 효소활성이 유지되는 것을 확인하였으며 이들 추출물들은 알코올 분해 촉진작용이 매우 우수함을 알았다.

• BMB Reports
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• 제33권4호
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• pp.317-320
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• 2000

The succinic semialdehyde dehydrogenase from bovine brain was inactivated by treatment with phenylglyoxal, a reagent that specifically modifies arginine residues. The inhibition at various phenylglyoxal concentrations shows pseudo-first-order kinetics with an apparent secondorder rate constant of 30 $M^{-1}min^{-1}$ for inactivation. Partial protection against inactivation was provided by the coenzyme $NAD^+$, but not by the substrate succinic semialdehyde. Spectrophotometric studies indicated that complete inactivation of the enzyme resulted from the binding of 2 mol phenylglyoxal per mol of enzyme. These results suggest that essential arginine residues, located at or near the coenzyme-binding site, are connected with the catalytic activity of brain succinic semialdehyde dehydrogenase.

• Journal of Applied Biological Chemistry
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• 제56권2호
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• pp.89-93
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• 2013

Gamma-aminobutyric acid (GABA) aminotransferase (gabT) and succinic semialdehyde dehydrogenase (gabD) genes from Pseudomonas fluorescens KCCM 12537 were cloned into a single pETDuet-1 vector and co-expressed in Escherichia coli BL21(DE3) simultaneously. The mixture of both enzymes, called GABase, is the key enzyme for the enzymatic analysis of GABA. The molecular mass of the GABA aminotransferase and succinic semialdehyde dehydrogenase were determined to be 52.8 and 46.7 kDa following computations performed with the pI/Mw program, respectively. The GABase activity between pH 6.0 and 9.0 for 24 h at $4^{\circ}C$ remained over 75%, but under pH 6.0 decreased rapidly. The GABase activity between 25 and $35^{\circ}C$ by the treatment at pH 8.6 for 30 min remained over 80%, but over $35^{\circ}C$ decreased rapidly. When the activity against GABA was defined as 100%, the purified GABase activity against 5-aminovaleric acid having a similar structure to GABA showed 47.7% and GABase activity against ${\beta}$-alanine, ${\varepsilon}$-amino-n-caproic acid, $_L$-ornithine, $_L$-lysine, and $_L$-aspartic acid showed between 0.3 to 2.3%. The GABA content was analyzed with this co-expressed GABase, compared with the other GABase which was available commercially. As a result, the content of GABA extracted from brown rice, dark brown rice, and black rice were $26.4{\pm}3.5$, $40.5{\pm}4.7$ and $94.7{\pm}9.3{\mu}g/g$, which were similar data of other GABase in the error ranges.

• 한국미생물·생명공학회지
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• 제19권1호
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• pp.31-36
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• 1991

Tn5의 kanamycin 저항성 유전자를 가진 pBEL1 plasmid와 C.glutamicum cryptic plasmid인 pSR1으로 7.5kb의 새로운 plasmid를 만든 후, 이를 pCE1301이라 명명하였다. 이 pCE1301은 PEG1301은 PEG-protoplast법으로 C.glutamicum을 형질전환하였을 때 효율이 약 $3.0\times 10^3$형질전환제/$\mu g$이었으며 SalI과 EcoRI 제한효소 절단부위가 1개씩이었다. 또 Km이 없는 배지에서 25세대까지 안정하게 유지되었으며 B.flavum, E.coli에서 복제되었다. 이 pCE1301을 이용하여 C.glutamicum 의 homoserine dehydrogenase 유전자를 cloning하였다.

• Journal of Microbiology and Biotechnology
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• 제24권12호
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• pp.1636-1643
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• 2014

3-Hydroxybutyryl-CoA dehydrogenase is an enzyme that catalyzes the second step in the biosynthesis of n-butanol from acetyl-CoA, in which acetoacetyl-CoA is reduced to 3-hydroxybutyryl-CoA. To understand the molecular mechanisms of n-butanol biosynthesis, we determined the crystal structure of 3-hydroxybutyryl-CoA dehydrogenase from Clostridium butyricum (CbHBD). The monomer structure of CbHBD exhibits a two-domain topology, with N- and C-terminal domains, and the dimerization of the enzyme was mostly constituted at the C-terminal domain. The mode of cofactor binding to CbHBD was elucidated by determining the crystal structure of the enzyme in complex with $NAD^+$. We also determined the enzyme's structure in complex with its acetoacetyl-CoA substrate, revealing that the adenosine diphosphate moiety was not highly stabilized compared with the remainder of the acetoacetyl-CoA molecule. Using this structural information, we performed a series of site-directed mutagenesis experiments on the enzyme, such as changing residues located near the substrate-binding site, and finally developed a highly efficient CbHBD K50A/K54A/L232Y triple mutant enzyme that exhibited approximately 5-fold higher enzyme activity than did the wild type. The increased enzyme activity of the mutant was confirmed by enzyme kinetic measurements. The highly efficient mutant enzyme should be useful for increasing the production rate of n-butanol.

• International Journal of Industrial Entomology and Biomaterials
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• 제35권1호
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• pp.22-29
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• 2017

Increased alcohol consumption is a burden on the world because it is associated with various health problems. However, the effects of silkworms on alcohol metabolism have not been studied yet. The hard-to-eat mature silkworms have become easier to ingest recently due to the development of technology, steam-lyophilising mature silkworm larvae. In this study, we investigated and compared the effects of SMSPs from three silkworm varieties, Baekokjam, Golden-silk and Yeonnokjam weaving white, golden, and light green cocoons on alcohol metabolism in vivo. Sprague-Dawley rats pretreated with three SMSPs (0.1 g/kg or 1 g/kg body weight) or normal diet (AIN-76A) for 2 weeks were subjected to intragastric administration of absolute ethanol (3 g/kg body weight, 3 h). Three SMSPs did not affect the final body weight and liver weight. All 3 SMSPs were effective to reduce the enzymes in alcohol metabolism, alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), and liver damage and enzymes involved in liver damage, aspartate aminotransferase (AST) and alanine aminotransferase (ALT). Among SMSP from 3 varieties of silkworm, preadministration of 1 g/kg Baekokjam SMSP showed the most effective suppressive effect on the activities of ADH, ALDH, AST and ALT. The Baekokjam SMSP contained higher amounts of beneficial amino acids than Golden-silk or Yeonnokjam SMSP. These results suggest that Baekokjam SMSP might be used as a new and promising candidate for improving alcohol metabolism and liver injury through promoting rapid alcohol metabolism.

• 한국식품영양과학회지
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• 제43권9호
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• pp.1363-1368
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• 2014

우리나라 남해안 연안에서 주로 양식되는 진주담치(국산 홍합)와 세계적으로 기능성이 잘 알려진 뉴질랜드 초록입 홍합의 열수 추출물을 제조하여 생리활성을 비교하였다. 모든 홍합 추출물들은 알코올대사와 관련한 alcohol dehydrogenase(ADH)와 acetaldehyde dehydrogenase(ALDH)의 활성을 상승시켰으며, 특히 ADH의 활성을 크게 향상시켰다. 국산 홍합의 육질 추출물은 초록입 홍합의 육질 추출물에 비하여 ADH 상승 효과는 비슷하였으며, ALDH 상승 효과는 약간 낮았다. DPPH 라디칼 소거능으로 조사한 항산화능은 국산 홍합의 육질 추출물이 초록입 홍합의 육질 추출물 보다 높았다. 항고혈압과 관련한 ACE 저해능의 경우에는 국산 홍합 육질 추출물이 초록입 홍합의 육질 추출물에 비해 10 mg/mL 농도에서는 낮았으나 20 mg/mL 이상의 농도에서는 유의차(P<0.05)를 보이지 않았다. 이상의 결과는 국내에서 양식되고 있는 홍합도 세계적으로 초록입 홍합에 못지않은 생리활성을 가지고 있음을 시사한다.

• Advances in Traditional Medicine
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• 제9권1호
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• pp.49-57
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• 2009

The antifertility activity of methanol extract of Cuscuta reflexa Roxb. stem (MECR) and Corchorus olitorius Linn. seed (MECO) were studied on male Swiss albino mice. The extracts were found to decrease sperm count, percentage of motile sperm and testosterone level in treated mice when compared with vehicle control after 17 days of treatment. The weight of gonads, epididymis were decreased whereas no significant changes of the body weight of mice were observed after methanol extract treatments. The fertility test showed 100% negative result in MECR and MECO treated mice at medium and high dose level of treatment. MECR and MECO in low (25 mg/kg and 15 mg/kg, respectively), medium (50 mg/kg and 20 mg/kg, respectively) and high (75 mg/kg and 25 mg/kg, respectively) dose level caused a simultaneous fall in testicular ${\Delta}5$-$3{\beta}$-hydroxy steroid dehydrogenase and glucose-6-phosphate dehydrogenase activities which are involved in testicular steroidogenesis. Total cholesterol and ascorbic acid content in testis were increased significantly in gonads. The activities of lactate dehydrogenase, malic dehydrogenase and ascorbic acid oxidase were reduced whereas that of carbonic anhydrase was increased significantly in the testis of MECR and MECO treated mice. All these observations indicate that the methanol extract of C. reflexa stem and C. olitorius seed produced antifertility activity in sexually matured male mice, which may be due to inhibition of gonadal steroidogenesis. This activity may be attributed due to the presence of flavonoids and steroids, respectively.

• BMB Reports
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• 제36권2호
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• pp.159-166
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• 2003

Cytoplasmic $\alpha$-glycerol-3-phosphate dehydrogenase from fruit-bat-breast muscle was purified by ion-exchange and affinity chromatography. The specific activity of the purified enzyme was approximately 120 units/mg of protein. The apparent molecular weight of the native enzyme, as determined by gel filtration on Sephadex G-100 was $59,500{\pm}650$ daltons; its subunit size was estimated to be $35,700{\pm}140$ by SDS-polyacrylamide gel electrophoresis. The true Michaelis-Menten constants for all substrates at pH 7.5 were $3.9{\pm}0.7\;mM$, $0.65{\pm}0.05\;mM$, $0.26{\pm}0.06\;mM$, and $0.005{\pm}0.0004\;mM$ for L-glycerol-3-phosphate, $NAD^+$, DHAP, and NADH, respectively. The true Michaelis-Menten constants at pH 10.0 were $2.30{\pm}0.21\;mM$ and $0.20{\pm}0.01\;mM$ for L-glycerol-3-phosphate and $NAD^+$, respectively. The turnover number, $k_{cat}$, of the forward reaction was $1.9{\pm}0.2{\times}10^4\;s^{-1}$. The treatment of the enzyme with 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) under denaturing conditions indicated that there were a total of eight cysteine residues, while only two of these residues were reactive towards DTNB in the native enzyme. The overall results of the in vitro experiments suggest that $\alpha$-glycerol-3-phosphate dehydrogenase of the fruit bat preferentially catalyses the reduction of dihydroxyacetone phosphate to glycerol-3-phosphate.