• Journal of Microbiology and Biotechnology
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• 제20권4호
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• pp.659-665
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• 2010

A cis-epoxysuccinate hydrolase (CESH) from Bordetella sp. BK-52 was purified 51.4-fold with a yield of 27.1% using ammonium sulfate precipitation, ionic exchange, hydrophobic interaction, molecular sieve chromatography and an additional anion-exchange chromatography. The CESH was stable in a broad range of temperature (up to $50^{\circ}C$) and pH (4.0-10.0) with optima of $40^{\circ}C$ and pH 6.5, respectively. It could be partially inhibited by EDTA-$Na_2$, $Ag^+$, SDS, and DTT, and slightly enhanced by $Ba^{2+}$ and $Ca^{2+}$. The enzyme exhibited high stereospecificity in D(-)-tartaric acid (enantiomeric excess value higher than 99%) with $K_m$ and $V_max$ values of 18.67 mM and $94.34\;{\mu}M$/min/mg for disodium cis-epoxysuccinate, respectively. The Bordetella sp. BK-52 CESH gene, which contained 885 nucleotides (open reading frame) encoding 294 amino acids with a molecular mass of about 32 kDa, was successfully overexpressed in Escherichia coli using a T7/lac promoter vector and the enzyme activity was increased 42-times compared with the original strain. It may be an industrial biocatalyst for the preparation of D(-)-tartaric acid.

• BMB Reports
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• 제38권1호
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• pp.49-57
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• 2005

Major Royal Jelly Protein cDNAs of Apis cerana (AcMRJP) were cloned and characterized. The open reading frames (ORFs) of the AcMRJP1 and AcMRJP2 genes were 1302 and 1392 nucleotides, encoding 433 and 463 amino acid residues, respectively. The sequence divergences between AcMRJP1 and AcMRJP2 and their corresponding protein families in A. mellifera were 0.0618 and 0.0934 at the nucleotide level and 0.0912 and 0.1438 at the protein level, respectively. Phylogenetic analysis supports the orthologous similarity between these proteins. The deduced amino acids indicated high essential amino acid contents of AcMRJP1 and AcMRJP2 (47.5 and 44.8%, respectively). The genomic organization of both AcMRJP1 and AcMRJP2 was determined. Both the AcMRJP1 (3663 bp) and AcMRJP2 (3963 bp) genes contained six exons and five introns, where all boundaries conformed to the GT/AG rule. AcMRJP1 and AcMRJP2 cDNAs were cloned into pET17b, and both the recombinant (r) AcMRJP1 (47.9 kDa) and rAcMRJP2 (51.7 kDa) were expressed in the insoluble form. Western blot analysis and N-terminal sequencing of the solubilized proteins revealed successful expression of rAcMRJP1 and rAcMRJP2 in vitro. The yields of the purified rAcMRJP1 and rAcMRJP2 were approximately 20 and 8mg protein per liter of the flask culture, respectively.

• Journal of Biosystems Engineering
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• 제34권1호
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• pp.50-56
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• 2009

Surface plasmon resonance (SPR) biosensor has been used to detect many biochemical reactions, because this label-free sensor has high sensitivity and rapid response. The reactions are monitored by refractive index changes of the SPR biosensor. Iprovalicarb is protective, curative, and eradicative systemic fungicide introduced by Bayer AG in 1999. It has potential far control of downy mildew infesting onion, cucumber, grape and melon, late blight infesting tomato and potato, and anthracnose infesting watermelon and pepper. It is strictly limited to the maximum residue limit. In this study, the applicability of a portable SPR biosensor (Spreeta, Texas instrument, TX, USA) to detect the iprovalicarb residue was examined. The sensor chip was adopted to detect the reaction of iprovalicarb to immobilized iprovalicarb-antibody. The binding of the iprovalicarb onto the biosensor surface was measured by change of the refractive index (RI). Characteristics of the sensor chip including specificity, sensitivity, stability, and reusability were analyzed. In calibration test for seven levels of iprovalicarb concentration (0.32 to 5,000 mg/L) with three replications, a Sigmoidal model with Hill function was obtained between relative RI value and the iprovalicarb concentration with R-square of 0.998. It took 30 minutes to complete a set of detecting assay with the SPR biosensor.

• Journal of Microbiology and Biotechnology
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• 제13권1호
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• pp.77-84
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• 2003

Chitinase was purified from an antagonistic bacterium Bacillus sp. 7079 by ammonium sulfate precipitation, QAE-Sephadex anion exchange chromatography, Sephadex G-100 gel filtration, and SP-Sephadex cation exchange chromatography. The molecula. weight of purified chitinase (PC-1) was approximately 66.5 kDa on SDS-PACE. PC-1 exhibited optimum pH and temperature of pH 7.5 and $45^{\circ}C$, respectively. More than $80\%$ of PC-1 was stable at pH 5.0 to 9.0, and more than $90\%$ at $40^{\circ}C$. $Fe^2+\;and\;Ca^2+$ inhibited the chitinase activity about $20\%$, and EDTA and p-CMB by about $30\%$, whereas $Ag^+$ inhibited the activity up to $65\%$. The $K_m$ value of PC-1 was 1.215 mg/ml with colloidal chitin as a substrate. We also investigated the effect of PC-1 treated chitin (PCTC) on the pro-inflammatory cytokine gene expression in macrophage RAW 264.7 cells. The expression of IL-$1{\alpha}$ and IL-$1{\beta}$ mRNA gene was investigated using reverse transcriptase polymerase chain reaction (RT-PCR). IL-$1{\alpha}$ and IL-$1{\beta}$ mRNA were induced by the treatment of PCTC and chitin only in RAW 264.7 cells. These expressions were induced as early as 2 h and sustained up to 24 h in RAW 264.7 cells. IL-$1{\alpha}$ and IL-$1{\beta}$ mRNA were more strongly expressed by the treatment of PCTC than chitin treatment alone in RAW 264.7 cells.

• Journal of Microbiology
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• 제35권2호
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• pp.109-116
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• 1997

An extracellular proteinase of Candida albicans was purified by a combination of 0~75% ammonium sulfate precipitation, DEAE Sepharose Fast Flow ion exchange chromatography, and Sephacryl S-200 HR molecular sieve chromatography. Its mlecular weight was approximately 41 kDa on SDS-PAGE and isoelectric point was 4.4. The enzyme was inhibited by pepstain A. Optimum enzyme activity ranged from pH 2.0 to 3.5 with its maximum at pH 2.5 and a temperature of 45$^{\circ}C$. The addition of divalent cations, $Ca^{2+}$, Zn$^{2+}$ and $Mg^{2+}$, resulted in no significant inhibition of enzymatic activity. However, some inhibitory effects were observed by Fe$^{2+}$, Ag$^{2+}$ and Cu$^{2+}$. With BSA as substrate, an apparent $K_m$ was determined to be 7$\times$10$^{-7}$ M and $K_i$, using pepstatin A as an inhibitor, was 8.05$\times$10$^{-8}$ M. N-terminal amino acid sequence was QAVPVTLXNEQ. Degradation of BSA and fibronectin was shown but not collagen, hemoglobin, immunoglobulin G, or lysozyme. The enzyme preferred peptides with Glu and Leu at the P$_1$ position, but the enzyme activity was highly reduced when the P$_2$ position was phe or pro. This enzyme showed antigenicity against sera of patients with candidiasis.

• Bulletin of the Korean Chemical Society
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• 제32권9호
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• pp.3223-3228
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• 2011

To quantify the presence of mercuric ions in aqueous solution, double-stranded DNA (dsDNA) of poly(dT) was employed using a light switch compound, $Ru(phen)_2(dppz)^{2+}$ (1) which is reported to intercalate into dsDNA of a right-handed B-form. Addition of mercuric ions induced the dehybridization of poly(dT)${\cdot}$poly(dA) duplexes to form a hairpin structure of poly(dT) at room temperature and the metal-to-ligand charge transfer emission derived from the intercalation of 1 was reduced due to the dehybridization of dsDNA. As the concentration of $Hg^{2+}$ was increased, the emission of 1 progressively decreased. This label-free emission method had a detection limit of 0.2 nM. Other metal ions, such as $K^+$, $Ag^+$, $Ca^{2+}$, $Mg^{2+}$, $Zn^{2+}$, $Mn^{2+}$, $Co^{2+}$, $Ni^{2+}$, $Cu^{2+}$, $Cd^{2+}$, $Cr^{3+}$, $Fe^{3+}$, had no significant effect on reducing emission. This emission method can differentiate matched and mismatched poly(dT) sequences based on the emission intensity of dsDNA.

• Carbon letters
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• 제8권4호
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• pp.274-279
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• 2007

In order to understand the breakthrough behaviour of iodine vapours on impregnated carbon systems, an active carbon, 80 CTC grade, $12{\times}30$ BSS particle size and $1104\;m^2/g$ surface area, was impregnated with metal salts such Cu, Cr, Ag, Mo and Zn, and an organic compound Triethylene diamine (TEDA) to prepare different carbon systems such as whetlerite, whetlerite/TEDA, whetlerite/KI/KOH and ASZMT. The prepared adsorbents along with active carbon were characterized for surface area and pore volume by $N_2$ adsorption at liquid nitrogen temperature. These carbon systems were compared for their CT (concentration X time) values at 12.73 to 53.05 cm/sec space velocities and 2 to 5 cm carbon column bed heights. The carbon column of 5.0 cm bed height and 1.0 cm diameter was found to be providing protection against iodine vapours up to 5.5 h at 3.712 mg/L iodine vapour concentration and 12.73 cm/sec space velocity. The study clearly indicated the adsorption capacities of carbon systems to be directly proportional to their surface area values. Dead layer with all the prepared carbon systems was found to be less than 2.0 cm indicating it to be minimum bed height to have protection against $I_2$ vapours. Effect of carbon bed height and flow rate was also studied. The active carbon showed maximum protection at all bed heights and flow rates in comparison to all other impregnated carbon systems, showing that only physical adsorption is responsible for the removal of iodine vapours.

• Journal of Microbiology and Biotechnology
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• 제20권11호
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• pp.1546-1554
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• 2010

A bacterial strain capable of producing extracellular ${\alpha}$-galactosidase was isolated from a sample of sugarcane industrial waste. Microbiological, physiological, and biochemical studies revealed that the isolate belonged to Bacillus sp. Furthermore, based on a 16S rDNA sequence analysis, the new isolate was identified as Bacillus megaterium VHM1. The production of ${\alpha}$-galactosidase was optimized based on various physical culture conditions. Guar gum and yeast extract acted as the best carbon and nitrogen sources, respectively. The optimum pH was 7.5 and the enzyme remained stable over a pH range of 5-9. The enzyme was optimally active at $55^{\circ}C$ and thermostable with a half-life of 120 min, yet lost 90% of its residual activity within 120 min at $60^{\circ}C$. One mM concentrations of $Ag^2$, $Cu^2$, and $Hg^{2+}$ strongly inhibited the ${\alpha}$-galactosidase, whereas the metal ions $Fe^2$, $Mn^{2+}$, and $Mg^{2+}$ had no effect on the ${\alpha}$-galactosidase activity, and $Zn^{2+}$, $Ni^{2+}$, and $Ca^{2+}$ reduced the enzyme activity slightly. When treated with the B. megaterium VHM1 enzyme, the flatulence-causing sugars in soymilk were completely hydrolyzed within 1.5 h.

• 자원환경지질
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• 제12권2호
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• pp.79-93
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• 1979

한반도 내에 부존되고 있는 중적광상들은 세 주요한 Metallousenetic Epoch (Pre-Cambrian E., Jurassic-early Cretaceous E., 및 late Cretaceous-early Tertiary E.)에 생성되었고 성인으로 열수작용과 접촉교대작용에 기인된다고 보고되었다. 이중 12개 광상(달성, 산내, 일광, 상동, 대화, 월악, 복수, 옥방, 쌍전, 홍성, 삼봉, 청양)을 본 연구의 대상으로 택하고 각 지역에서 채취한 표품중 모두 25개 시료에 대하여 지화학적 실험을 수행하였다. 즉 각 분쇄된 시료 (-80+120mesh)는 super panner, 중액, isodynamic separator, UV lamp를 이용한 물리적인 처리과정을 거쳐 최종으로 입체현미경 하에서 단체분리된 후 Jarrell-Ash 1.5m Grating Spectrometer를 이용하여 본 연구대상 시료로 개발한 Spectrochemical method (Carrier: NaCl, Internal standard: $La_2O_3$)로 정량 분석되었다. 본 연구에서 밝혀낸 국내 회중석 광내에 함유되는 희유 윈소의 종류는 모두 Al, Bi, Fe, Si, Mn, Pb, Mg, Sn, Mo, Cu, Sr, Cr, Y, Ag, Ti, Ni, As, Yb의 18개 원소이며, 이들은 각각의 절대적 및 상대적인 함량과 각 광화작용의 시기, 광상의 성인, 모암, 각 원소의 mobility 그리고 회중석광의 형광색 및 육안색과 비교 검토되었다. 본 연구의 결과로 회중석광이 정출되는 과정에서 유사한 지질환경의 물리화학적인 여건이 이루어지면 그 내에 함유되는 몇 성분원소들의 화학적인 특성은 매우 유사성을 보여주는 typochemical habit를 가짐을 알 수 있었다. 본 연구에서 밝혀진 한국산 회중석 광내의 희유성분(稀有成分)으로서 Y, Mn, Sr 원소들의 Geochemical mobility의 특성(特性)은 앞으로 회중석광의 심도탐광을 위하여 고려되어야하고, 더욱 연구개발되어야 할 것이다. 또한 상기 회중석 광내의 물리적 및 화학적으로 결합되어 있는 불순물의 희유성분들은 선광 제련 파정에서 금속성분의 추출농집공정(工程)에 유용한 자료가 될 것이다.

• 한국미생물·생명공학회지
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• 제20권2호
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• pp.150-155
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• 1992

토양 분리 균주로부터 얻은 배양액을 20-80 황산암모늄으로 분획한 후 세포의 cytosine deaminase의 성질을 검토하였다. 이 효소는 cytosine과 5-fluorocytosine(5-FC)에 기질 특이성을 가짐으로서 각각 uracil과 5-fluorouracil(5-FU)로 전환을 촉매하였다. 효소안정성에 대한 온도와 보존기간은 tris-HC1 완충용액에서 검토한 결과 최적온도는 $50^{\circ}C$ 부근이하에서 90 이상의 잔존활성을, 보존시간은 4일정도에서 80 이상의 잔존활성을 유지하였다. 최대 pH 8.0과 $37^{\circ}C$의 온도에서 나타났다. 활성에 적당한 pH는 8.0~8.5였고, 온도는 37~$45^{\circ}C$의 범위였다.