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방선균에 의해 생산된 항 MRSA 항생물질 AM3 의 구조 연구
임융호(Yoong Ho Lim),장준환(Jun Hwan Chang),김종훈(Jong Hoon Kim),서정우(Jung Woo Suh),정재경(Jae Kyung Jung),이철훈(Chul Hoon Lee) 한국응용생명화학회 1995 Applied Biological Chemistry (Appl Biol Chem) Vol.38 No.6
In order to find a potent anti-methicillin resistant Staphylococcus aureus (MRSA) antibiotic, actinomycetes isolated from the samples collected in Korean marine silt were screened. From the culture broth of the isolated Streptamyces strain AM045, a substance showing excellent biological activity against MRSA was found, isolated and named AM3. The compound showed strong activities against MRSA, S. epideridis, E. faecium and E. faecalis, which were better than those of vancomycin and teicoplanin. Unfortunately, AM3 was identified as Actinomycin V. However, this paper reports the three dimensional study of AM3 based on high resolution nmr and Computer Aided Molecular Modeling(CAMM), and the fact that the structure of the pentapeptide lactone ring with oxo-proline in chloroform solution does not have $quot;C conformation$quot; any more.
Streptomyces coelicolor A3(2)로 부터 $\beta$-Glucosidase 유전자 클로닝 및 재조합 효소의 특성
김재영,김봉규,이용섭,강창수,안중훈,임융호,Kim, Jae-Young,Kim, Bong-Kyu,Yi, Yong-Sub,Kang, Chang-Soo,Ahn, Joong-Hoon,Lim, Yoong-Ho 한국미생물·생명공학회 2009 한국미생물·생명공학회지 Vol.37 No.2
The $\beta$-glucosidase gene from Streptomyces coelicolor A3(2) was cloned and expressed in Escherichia coli. The ORF consisted of 1377 nucleotides encoding 51 kDa in a predicted molecular weight. Effects of pH indicated that the $\beta$-glucosidase showed similar activity using $\alpha$-pNPG($\rho$-nitrophenyl-$\alpha$-D-glucopyranoside), $\beta$-pNPG($\rho$-nitrophenyl-$\beta$-D-glucopyranoside), and $\beta$-pNPF($\rho$-nitrophenyl-$\beta$-D-fucopyranoside) at range of pH 3 to 10, and high activity using $\beta$-pNPGA ($\rho$-nitrophenyl-$\beta$-D-galactopyranoside) from pH 5 to 10, especially, 3.3 times higher activity at pH 9. Effects of temperature indicated that the $\beta$-glucosidase showed low activity using $\alpha$-pNPG, $\beta$-pNPG, and $\beta$-pNPF from $20^{\circ}C$ to $70^{\circ}C$, and increased activity using $\beta$-pNPGA from $30^{\circ}C$ to $50^{\circ}C$, 1.8 times higher activity at $50^{\circ}C$ than at $30^{\circ}C$. According to activity determination of other substrates, the enzyme was active on daidzin, genistin, and glycitin, inactive on esculin and apigenin-7-glucose. The EDTA and DTT as reducing agents inhibited $\beta$-glucosidase activity, but SDS and mercaptoethanol did not inhibit. Monovalent or divalent metal ions such as $MnSO_4$, $CaCl_2$, KCl, and $MgSO_4$ did not inhibited $\beta$-glucosidase activity. $CuSO_4$ and NaCl showed low inhibition, and $ZnSO_4$ inhibited 3.3 times higher than control. Streptomyces coelicolor A3(2)의 $\beta$-glucosidase 유전자를 분리하여 대장균에서 발현하여 특성을 조사하였다. 최적 활성을 나타내는 온도는 pH 5에서는 $20^{\circ}C$, pH 6에서는 $60^{\circ}C$에서 높은 활성을 나타냈다. pH에 따른 활성은 pH 3 이하와 pH 9 이상의 범위에서는 낮은 활성을 나타냈으며 pH 7에서 가장 높은 활성을 나타냈다. $\alpha$-pNPG($\rho$-nitrophenyl-$\alpha$-D-glucopyranoside), $\beta$-pNPG ($\rho$-nitrophenyl-$\beta$-D-glucopyranoside), $\beta$-pNPF($\\rho$-nitrophenyl-$\beta$-D-fucopyranoside)는 pH 3-10까지 비슷한 활성을 나타냈으며, $\alpha$-pNPG가 pH 7에서 다소 높은 활성을 보였다. $\beta$-pNPGA는 pH 5-9까지 높은 활성을 나타냈으며, 특히 pH 9에서 3배 이상의 높은 활성을 나타냈다. 기질 $\alpha$-pNPG, $\beta$-pNPG, $\beta$-pNPF의 온도에 따른 활성변화는 $\beta$-pNPF의 활성이 $60^{\circ}C$에서 증가하였고, $\beta$-pNPGA는 $30-50^{\circ}C$까지 활성이 증가하여 $50^{\circ}C$에서 최대활성을 나타내었다. 당화 flavonoid를 이용한 기질특이성의 상대활성은 daidzin, glycitin, genistin, 순으로 나타났으며 esculin과 apigenin-7-glucose는 기질로 사용하지 않았다. $\beta$-Glucosidase 활성은 EDTA, DTT에 의해 억제되었으며, $MnSO_4$, $CaCl_2$, KCl, $MgSO_4$에 의해 증가하였고, 특히 Mn이온에 의해 증가하였다. $CuSO_4$, NaCl에 의해 효소활성이 저해되었으며, 특히 $ZnSO_4$의 경우 효소활성이 강하게 억제되었다.
식품 및 환경, 기타 한국 전통젓갈에서 분리한 Bacillus subtilis JKK238 균주 유래 세 종류 Lipopeptide의 분리 및 특성
윤상홍 ( Sang Hong Yoon ),김정봉 ( Jung Bong Kim ),임융호 ( Yoong Ho Lim ),홍성렬 ( Seong Ryeul Hong ),송재경 ( Jae Kyeung Song ),김삼선 ( Sam Sun Kim ),권순우 ( Soon Wo Kwon ),박인철 ( In Cheol Park ),김수진 ( Soo Jin Kim ),여윤 한국미생물생명공학회 2005 한국미생물·생명공학회지 Vol.33 No.4
수종의 식물수출물의 항산화 및 Melanin 합성 억제효과
김재영 ( Jae Young Kim ),이진영 ( Jin Young Lee ),이위영 ( Wi Young Lee ),이용섭 ( Yong Sub Yi ),임융호 ( Yoong Ho Lim ) 한국미생물생명공학회 ( 구 한국산업미생물학회 ) 2010 한국미생물·생명공학회지 Vol.38 No.4
본 연구에서는 식물 추출물을 이용하여 항산화 활성 및 tyrosinase 활성 억제 효과를 측정하였다. 식물 추출물의 폴리페놀물질의 총 함량은 Acer psedo-siebolianum의 추출물이 16.4 mg/g로 가장 높은 추출량을 나타내었다. 항산화활성 측정에서는 Acer ginnala 에서 IC50값으로 21.3 μg/mL으로 가장 좋은 활성을 나타내었다. 반면에 L-DOPA를 기질로 하여 mushroom tyrosinase의 활성 억제측정에서는 Distylum racemosum 1,000 mg에서 49.1%로 다른 추출물에 비하여 상대적으로 높은 활성을 나타내었다. Tyrosinase 활성 억제력이 가장 높은 D. racemosum의 추출물을 이용하여 ethanol 분획과 ethyl acetate 분획으로 분리하여, 이중 D. racemosum의 ethanol 분획에서 항산화 활성 IC50 값은 0.9 μg/mL, tyrosinase 활성억제는 IC50값이 118.1 μg/mL로 ethyl actate 분획보다 높은 활성을 나타내었다. 또한 ethanol 분획을 이용하여 B16/F1 melanoma cell에서는 60 μg/mL까지는 세포독성을 나타내지 않았으며 80 μg/mL의 농도에서 약간의 세포독성을 나타내었다. 에탄올 분획을 이용한 세포내 melanin 색소의 생산억제 IC50값은 75.4μg/mL로 분석되었다. 이러한 결과로 D. racemosum의 에탄올 추출물이 B16/F1 melanoma cell세포의 melanin색소합성대사에 관여하여 색소합성을 저해하는 것으로 보인다. Plants extracts are good resources to find functional compounds for human health. The following eight plants were collected and total phenolic contents were determined. Acer psedo-siebolianum showed the highest phenolic contents, 16.4 mg/g, whereas Cercidiphyllum japonica showed the lowest contents, 1.9 mg/g. The DPPH free radical scavenging capacities of the plant extracts showed high activity in following order : Acer ginnala (21.3 μg/mL) > Cornus walteri (23.9 μg/mL) > Distylum racemosum (29.2 μg/mL) > Castanopsis cuspidata var. Thunbergii (31.7 μg/mL) > Acer psedo-siebolianum (34.6 μg/mL) > Thuijopsis dolabrata cv. Aurea (53.1 μg/mL) > Cercidiphyllum Japonica (115.2 μg/mL). Also the mushroom tyrosinase inhibitory activities of total extracts were determined at different concentration. D. racemosum extract showed highest (49.1% at 1,000 mg) in inhibitory activity than other seven extracts. The ethanol fraction (IC50 value: 118.1 μg/mL) from D. racemosum showed more inhibitory activity than ethyl acetate fraction (IC50 value: 203 μg/mL). The ethanol fraction on showed no significant cytotoxicity in B16/F1 cells line up to 60 μg/mL. Over 80 μg/mL of ethanol fraction showed cytotoxicity in B16/F1 cells. The melanin contents of cells were significantly attenuated by ethanol fraction in a dose-dependent manner. The IC50 value of ethanol fraction was 75.4 μg/mL.
유전자조작, 균주분리 Streptomyces coelicolor A3(2)로 부터 β-Glucosidase 유전자 클로닝 및 재조합 효소의 특성
김재영 ( Jae Young Kim ),김봉규 ( Bong Kyu Kim ),이용섭 ( Yong Sub Yi ),강창수 ( Chang Soo Kang ),안중훈 ( Joong Hoon Ahn ),임융호 ( Yoong Ho Lim ) 한국미생물생명공학회 2009 한국미생물·생명공학회지 Vol.37 No.2
The β-glucosidase gene from Streptomyces coelicolor A3(2) was cloned and expressed in Escherichia coli. The ORF consisted of 1377 nucleotides encoding 51 kDa in a predicted molecular weight. Effects of pH indicated that the β-glucosidase showed similar activity using α-pNPG(ρ-nitrophenyl-α-D-glucopyranoside), β-pNPG(ρ-nitrophenyl-β-D-glucopyranoside), and β-pNPF(ρ-nitrophenyl-β-D-fucopyranoside) at range of pH 3 to 10, and high activity using β-pNPGA (ρ-nitrophenyl-β-D-galactopyranoside) from pH 5 to 10, especially, 3.3 times higher activity at pH 9. Effects of temperature indicated that the β-glucosidase showed low activity using α-pNPG, β-pNPG, and β-pNPF from 20℃ to 70℃, and increased activity using β-pNPGA from 30℃ to 50℃, 1.8 times higher activity at 50℃ than at 30℃. According to activity determination of other substrates, the enzyme was active on daidzin, genistin, and glycitin, inactive on esculin and apigenin-7-glucose. The EDTA and DTT as reducing agents inhibited β-glucosidase activity, but SDS and mercaptoethanol did not inhibit. Monovalent or divalent metal ions such as MnSO4, CaCl2, KCl, and MgSO4 did not inhibited β-glucosidase activity. CuSO4 and NaCl showed low inhibition, and ZnSO4 inhibited 3.3 times higher than control.