• Applied Biological Chemistry
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• 제49권4호
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• pp.315-321
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• 2006

유기인계나 카바메이트 살충제는 신경전달계 효소인 Acetylcholinesterase(AchE)를 저해하므로 농산물중에서 농약의 잔류여부 확인이 가능하다. 이러한 바이오센서는 환경 관리를 위한 민감한 센서로 사용될 수 있으며, 최근에는 AchE를 이용한 잔류농약을 측정할 수 있는 바이오센서 기술이 주목을 받고 있다. 우리는 최근에 감광성고분자물질인 고정화물질을 이용하여 광기교화반응에 의해 AchE를 막에 고정화시켜 잔류농약을 측정할 수 있는 바이오센서를 개발하였다. 고정화시킨 효소는 $4^{\circ}C$에서 6개월 동안 안정하였으며, 유기인계와 카바메이트계의 농약에 대한 반응에서도 기존의 용액형태의 효소보다 민감한 반응을 보였다. 또한 본 연구개발로 제작된 바이오센서에 의한 잔류농약 측정법은 용액형태의 효소에 의한 잔류농약 측정법보다 신속 간편하여 5분 안에 잔류농약에 대한 효소의 억제율을 측정할 수 있었으며, chlorpyrifos, cabaryl, carbofuran methidathion에 대해 농약의 검출 능력이 정밀분석법에 의한 검출한계에 근접한 것으로 나타났다

• Journal of Microbiology and Biotechnology
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• 제13권3호
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• pp.373-377
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• 2003

Various groups of industrial and agricultural pollutants (heavy metal ions, cyanides, and pesticides) can be detected by enzymes. Since heavy metal ions inhibit urease, cyanides inhibit peroxidase, organophosphorus and carbamate pesticides inhibit butyrylcholinesterase, these enzymes were co-immobilized into a bovine serum albumin gel on the surface of an ion-sensitive field effect transistor to create a bioprobe that is sensitive to the compounds mentioned above. The sensitivity of the present sensor towards KCN corresponded to $1\;\mu\textrm{M}$ with 1 min of incubation time. The detection limits for Hg(II) ions and the pesticide carbofuran were 0.1 and $0.5\;\mu\textrm{M}$, respectively, when a 10 min sensor incubation time in contaminated samples was chosen. The total time for determining the concentrations of all species mentioned did not exceed 20 min.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2003년도 제10차 국제학술회의 및 추계정기 학술발표회
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• pp.17-17
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• 2003

Enzymes involved in catecholamine synthesis are present in the highest concentration in the adrenal medulla, however they were found also in other, mainly nervous tissues. Increased transcription of genes for catecholamine biosynthetic enzymes is an important mechanism to increase the capacity for epineprine/norepinephrine biosynthesis with stress. Gastrodia elata(Chinese name: Tienma), are very important Chinese herbal medicines used for the medical treatment of headaches, migraine, dizziness, epilepsy, rheumatism, neuralgia, paralysis and other neuralgic and nervous disorders. Immobilize stressed rat markedly increased tyrosine hydroxylase (TH) mRNA and dopamine-${\beta}$-hydroxylase (DBH) mRNA transcriptior level more than control group. But treated Gastrodia elata extracts in immobilized stressed rat slightly increased TH mRNA and DBH mRNA transcription level more than normal group. In addition, we are obtained identical results in PC12 cell line. Decrease of transcription level of TH mRNA and DBH mRNA is indicating that Gastrodia elata have a anti-stress effects which decrease the transcription level of TH and DBH mRNA on catecholamine biosynthesis pathway.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2003년도 심포지엄
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• pp.37-37
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• 2003

Enzymes involved in catecholamine synthesis are present in the highest concentration in the adrenal medulla, however they were found also in other, mainly nervous tissues. Increased transcription of genes for catecholamine biosynthetic enzymes is an important mechanism to increase the capacity for epineprine/norepinephrine biosynthesis with stress. Gastrodia elata(Chinese name: Tienma), are very important Chinese herbal medicines used for the medical treatment of headaches, migraine, dizziness, epilepsy, rheumatism, neuralgia, paralysis and other neuralgic and nervous disorders. Immobilize stressed rat markedly increased tyrosine hydroxylase (TH) mRNA and dopamine-$\beta$-hydroxylase (DBH) mRNA transcription level more than control group. But treated Gastrodia elata extracts in immobilized stressed rat slightly increased TH mRNA and DBH mRNA transcription level more than normal group. In addition, we are obtained identical results in PC12 cell line. Decrease of transcription level of TH mRNA and DBH mRNA is indicating that Gastrodia elata have a anti-stress effects which decrease the transcription level of TH and DBH mRNA on catecholamine biosynthesis pathway.

• KSBB Journal
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• 제5권3호
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• pp.279-291
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• 1990

Chitin을 35 mesh로 분쇄하여 강산과 강알카리로 처리하여 CHITA와 CHITB를 만들고 여기에 glutaraldehyde를 작용시켜 $\beta$-glucosidase를 고정하여 CHITA-Gase 및 CHITB-Gase를 제조하였다. 이 두 종류의 고정화 효소를 기질 p-nitropheol-$\beta$-D-gliucopyranoside(PNG)과 회분식 반응기, 연속 흐름 반응기 및 플러그 흐름 반응기에서 반응시켜 최적 온도, 최적 pH, 반응 속도 상수, 물질 전달 계수, 효율 인자 및 효소 불활성화 속도 등을 구하여 반응기별 효율을 검토하였다. 반응 최적 온도는 세가지의 반응기 모두 5$0^{\circ}C$였으며, 최적 pH는 플러그 흐름 반응기에서는 Nat-Gase와 같이 pH5.0이었고 회분식 반응기 및 연속 흐름 반응기에서는 최적 pH의 이동이 일어나 pH6.0으로 이었다. 반응기의 최적 조건에서 km값은 회분식 반응기에서 CHITB-Gase$1.725$\times$10^-^5M/1$가 CHITA-Gase($1.725$\times$10^-^5M/1$)보다 작았으며, 연속 흐름 반응기 및 플러그 흐름 반응기에서는 유속 증가에 따라 Km'치가 감소하는 경향을 보였고, CHITB-Gase가 CHITB-Gase보다 더 작았다. $V^m^a^x'$값은 회분식 반응기, 연속 흐름 반응기, 플러그 흐름 반응기에서 모두 CHITA-Gase가 CHITB-Gase보다 높은 것으로 나타났다. 그리고, 물질전달계수 및 효율인자, 효소 불활성화 속도등의 값은 환경은 CHITB-Gase의 것이 나은 것으로 나타났다. 이들의 결과에서 CHITA-Gase 및 CHITB-Gase는 기질과의 반응 환경이 좋으므로 chitind은 $\beta$-glucosidase의 좋은 지지체라고 판단되어 공업적 응용이 기대된다.

• 분석과학
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• 제23권2호
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• pp.172-178
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• 2010

Glutathione S-transferase는 식물의 해독작용에 중추적인 역할을 하는 화학 효소이다. 본 연구에서는 제초제 검출 키트 개발에 응용을 위하여 식물 해독작용에 중추적인 역할을 하는 glutathione Stransferase의 고정화 방법을 연구하였다. Chloroacetanilide계 제초제에 높은 효소 활성을 보이는 벼 유래 OsGSTF3에 공유결합을 통한 polystyrene-alkylamine 비드와 리간드결합을 통한 agarose-aminoalkyl 비드,포괄법을 통한 Na-alginate 비드를 이용하여 고정화를 실시하였다. 정제된 OsGSTF3 10 mg을 사용하여 고정화 하였을 때 0.62 mg/g 비드로 polystyrene-alkylamine에 가장 효율적으로 고정화 되었다. 고정화 된 OsGSTF3의 효소 활성은 야생형의 30%를 나타내었으며, 재사용에 의한 효소활성 측정시 3회 까지 처음 활성의 80% 이상을 유지하였다.

• Journal of Microbiology and Biotechnology
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• 제22권6호
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• pp.818-825
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• 2012

Keratinases are exciting keratin-degrading enzymes; however, there have been relatively few studies on their immobilization. A keratinolytic protease from Chryseobacterium sp. kr6 was purified and its partial sequence determined using mass spectrometry. No significant homology to other microbial peptides in the NCBI database was observed. Certain parameters for immobilization of the purified keratinase on chitosan beads were investigated. The production of the chitosan beads was optimized using factorial design and surface response techniques. The optimum chitosan bead production for protease immobilization was a 20 g/l chitosan solution in acetic acid [1.5% (v/v)], glutaraldehyde ranging from 34 g to 56 g/l, and an activation time between 6 and 10 h. Under these conditions, above 80% of the enzyme was immobilized on the support. The behavior of the keratinase loading on the chitosan beads surface was well described using the Langmuir model. The maximum capacity of the support ($q_m$) and dissociation constant ($K_d$) were estimated as 58.8 U/g and 0.245 U/ml, respectively. The thermal stability of the immobilized enzyme was also improved around 2-fold, when compared with that of the free enzyme, after 30 min at $65^{\circ}C$. In addition, the activity of the immobilized enzyme remained at 63.4% after it was reused five times. Thus, the immobilized enzyme exhibited an improved thermal stability and remained active after several uses.

• Applied Biological Chemistry
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• 제35권5호
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• pp.410-414
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• 1992

Streptomyces sp. S56 endoinulase를 DEAE-cellulose에 고정화하였고 고정화 효소의 일반 특성을 유리효소와 비교하였으며 돼지감자 추출물 및 inulin의 고정화 효소에 의한 가수분해 조건을 조사하였다. 고정화 효소의 최적 작용 pH는 $4.5{\sim}5.5$로 유리효소보다 효소작용 pH가 산성으로 이동되었으며 작용 안정 pH는 6으로 유리효소와 동일하나 전반적으로 고정화에 의하여 pH 안정성은 저하되었다. 반면 온도 안정성은 고정화에 의하여 크게 증가되어 $50^{\circ}C$, 1시간 처리에서도 70%, 이상의 활성을 가지고 있었으며 또한 최적온도도 유리효소보다 높게 나타났다. 기질친화성은 고정화에 의하여 5배 정도 낮아지나 반응 최고속도는 반대로 2배 이상 증가되었다. 돼지감자 추출물 및 inulin은 고정화 효소에 의하여 48시간 내에 각각 총당의 63%, 78%를 가수분해, fructose 및 inulobiose를 주로 생산하였다.

• Journal of Microbiology and Biotechnology
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• 제16권5호
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• pp.678-682
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• 2006

To remove nitrate from wastewater, a novel bioelectrochemical denitrification system is introduced. In this proposed system, biological reactions are coupled with reactions on the electrode, whereby the electrons are transferred to the bacterial enzymes via a mediator as an electron carrier. The denitrification reaction was achieved with permeabilized Ochrobactrum anthropi SY509 containing denitrifying enzymes, such as nitrate reductase, nitrite reductase, and nitrous oxide reductase, and methyl viologen was used as the mediator. The electron transfer from the electrode to the enzymes in the bacterial cells was confirmed using cyclic voltammetry. A high removal efficiency of nitrate was achieved when the bioelectrochemical system was used with the permeabilized cells. Furthermore, when the permeabilized cells were immobilized to a graphite felt electrode using a calcium alginate matrix containing graphite powder, a high removal efficiency was achieved (4.38 nmol/min mg cell) that was comparable to the result when using the free permeabilized cells.

• BMB Reports
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• 제31권5호
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• pp.519-523
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• 1998

We have previously reported that anti-glucose-6-phosphate dehydrogenase (G6PD) serum raised against native G6PD (nG6PD) enzyme recognized nG6PD antigen poorly in competitive enzyme-linked immunosorbent assay (ELISA) (Kim, 1997). In the present study, we investigated whether anti-G6PD serum raised against nG6PD can react with denatured G6PD effectively in competitive ELISA. We used partially active G6PD (paG6PD) by repeated freeze-thawing or SDS-denatured G6PD (SDS-G6PD) as both immobilized and soluble antigens, and anti-G6PD serum raised against nG6PD for competitive ELISA. The polystyrene cuvettes coated with either paG6PD or SDS-G6PD were challenged with a mixture of a limiting amount of anti-G6PD serum and various doses of paG6PD or SDS-G6PD as competitors, followed by incubation with alkaline phosphatase-anti-IgG conjugate. The competitive ELISA with paG6PD or SDS-G6PD antigen exhibited the sigmoidal dose-response curve characteristic of competition immunoassays. Furthermore, Triton-denatured G6PD (Triton-G6PD) was used in competitive ELISA. The paG6PD, SDS-G6PD, or Triton-G6PD used as competitors increased the inhibition of antibody binding to immobilized either of nG6PD or denatured G6PD compared with nG6PD competitor. The inhibition by denatured G6PD competitors was more pronounced at high competitor concentrations than at low counterparts. We conclude that anti-G6PD serum raised against nG6PD can effectively react with denatured G6PD in competitive ELISA and that our anti-G6PD serum recognizes denatured enzymes better than active enzymes.