곰팡이균 , 이스트 , 스트렙토마이쎄테스와 박테리아의 나이트릴기 가수분해 효소들에 관한 연구 α - 아미노 페닐아세토나이트릴의 나이트릴기를 가수분해시키어 페닐글라이신을 얻는 생화학적 방법의 개발

구양모 ( Yang Mo Goo ) 생화학분자생물학회 1986 BMB Reports Vol.19 No.1

When α-aminophenylacetonitrile was fed to fungi, yeast, Streptomycetes, or bacteria growing in nutrient and Theriault`s media, most of them were found to accumulate phenylglycine in the media. Some of them, A. unguis, A. oryzae, P. purpurogenum, B. spectabilis, B. subtilis, E. coli, and R. rubra cultured in the nutrient media, and P. varioti, B. spectabilis, G. subo.xydans, R. glutinis, and R. toruloides cultured in the Theriault`s media showed accumulation of phenylglycine in a large quantity. It was found that, when the organisms were examined by a cell contact method for the converion of the substrate to phenylglycine by incubation of the substrate with harvested cells in a buffer for 5 hrs, P. digitatum, B. spectabilis, and R. toruloides grown in the Theriault`s media and B. spectabilis, R. glutinis, and R. toruloides grown in the nutrient medium showed strong nitrite group hydrolyzing enzymes` activity when they were induced by the substrate. Most of them did not show any conversion of the substrate to phenylglycine or phenylglycineamide without preinduction except A. gigantus, A. flaviple, A. panamensis, P. varioti, E. coli, P. vulgaris, P. aeruginosa, and R. toruloides. Also, no microorganisms examined in this experiment showed production of phenylglycineamide before or after pre-induction in the culture or in the cell contact media.

항생물질 생산 토양 Actinomycetes 균주 선별과 항생물질 생산특성 조사

구양모(Yang Mo Goo),이윤영(Youn Young Lee),정연숙(Youn Sook Chung),이영복(Young Bok Lee),조영애(Young Ae Joe),조희영(Hee Yeong Cho),고영선(Young Sun Koh),이창훈(Chang Hoon Lee) 대한약학회 1991 약학회지 Vol.35 No.3

Selective culture of actinomycetes from soil microbes and their antibiotic producing characters by agar-disk method were examined. Some of the organisms which produced antibiotics on agar disk did not produce antibiotics in liquid culture. Further examination indicated that production of antibiotic was dependent on the composition of medium. Many streptomycestes produced antibacterial substances in tryptic soy broth but others produced antifungal antibiotics in V-8 broth. Production of antibacterial substances by Streptomyces sp. was also dependent on the medium composition.

Nitrile Group Hydrolyzing Enzymes in Fungi, Yeast, Streptomycetes, and Bacteria; Biochemical Hydrolysis of the Nitrile Group in $\alpha$-Aminophenylacetonitrile to Phenylglycine

구양모,Goo, Yang-Mo 생화학분자생물학회 1986 한국생화학회지 Vol.19 No.1

$\alpha$-아미노페닐아세토나이트릴을 곰팡이균, 이스트, 스트렙토마이세트스 또는 박테리아를 영양 또는 쎄리올트의 배지에서 키우면서 넣어주었을 때 대부분의 미생물들은 페닐글라이신을 배지에 누적하는 것이 발견되었다. 이들 몇몇, 특히 영양배지에 키운 A. unguis, A. oryzae P. purpurogenum, B. spectabilis, B. subtilis, E. coli, 그리고 R. rubra와 세리올트 배지에서 키운 P. varioti, B. spectabilis, G. suboxydans, R. glutinis와 R. toruloides들은 배지에 페닐글라이신을 대량, 누적시키는 것으로 나타났다. 이들 균준들을 기질과 함께 키워 수확하여 버퍼에 넣어 기질과 함께 5시간 동안 인큐베이션시키었을 때 기질이 페닐글라이신으로 변화되는 것을 세포접촉법에 의하여 조사하였을 때 쎄리올트 배지에 키운 P. digitatum, B. spectabilis와 R. toruloides와 영양배지에 키운 B. spectabilis, R. glutinis 와 R. toruloides들이 나이트릴기 가순분해 효소환성을 강하게 나타났다. 이들의 대부분은 기질에 의하여 유도되지 않을 때는 기질을 페닐글라이신이나 페닌글라이신아미드로 변화시키는 효소활성은 나타나지 않았는데 A. gigantus, A. flaviple, A. panamensis, P. varioti, E. coli, P. vulgaris, P. aeruginosa와 R. toruloides 경우는 효소활성이 나타났다. 또한 세포접촉법에 의하여 또는 기질을 성장배지에 넣어 주었을 때 이 실험에서 조사한 어떤 미생물도 효소 유도전이나 유도후에 페닐글라이신아미드 생성은 관찰되지 않았다. When $\alpha$-aminophenylacetonitrile was fed to fungi, yeast, Streptomycetes, or bacteria growing in nutrient and Theriault's media, most of them were found to accumulate phenylglycine in the media. Some of them, A. unguis, A. oryzae, P. purpurogenum, B. spectabilis, B. subtilis, E. coli, and R. rubra cultured in the nutrient media, and P. varioti, B. spectabilis, G. suboxydans, R. glutinis, and R. toruloides cultured in the Theriault's media showed accumulation of phenylglycine in a large quantity. It was found that, when the organisms were examined by a cell contact method for the converion of the substrate to phenylglycine by' incubation of the substrate with harvested cells in a buffer for 5 hrs, P. digitatum, B. spectabilis, and R. toruloides grown in the Theriault's media and B. spectabilis, R. glutinis, and R. toruloides grown in the nutrient medium showed strong nitrile group hydrolyzing enzymes' activity when they were induced by the substrate. Most of them did not show any conversion of the substrate to phenylglycine or phenylglycineamide without preinduction except A. gigantus, A. flaviple, A. panamensis, P. varioti, E. coli, P. vulgaris, P. aeruginosa, and R. toruloides. Also, no microorganisms examined in this experiment show,ed production of phenylglycineamide before or after pre-induction in the culture or in the cell contact media.

Aminoglycoside Antibiotic Modifying Enzymes in Aminoglycoside Producing Streptomycetes and Micromonospora spp.

구양모,최석례,김공환,임번삼,이새배,Goo, Yang-Mo,Choi, Seok-Rye,Kim, Kong-Hwan,Lim, Bun-Sam,Lee, Sae-Bae 생화학분자생물학회 1989 한국생화학회지 Vol.22 No.4

Streptomycin을 생성하는 7종의 S. griseus와 1종의 S. galbus의 세포 추출물은 streptomycin을 phosphorylation하여 비활성화 하였다. Phosphorylation에 의한 streptomycin의 비활성화는 모든 streptomycin 생성균주에 공통적으로 존재하는 것으로 생각된다. Neomycin과 paromomycin을 생성하는 S. fradiaea와 S. rimosus forma paromomycinus의 세포추출물은 phosphorylation과 acetylation에 의하여 neomycin을 비활성화하였다. Kanamycin을 생성하는 S. kanamyceticus IFO 13414의 세포추출물도 neomycin을 phosphoryltion 또는 acetylabon시켜 비활성화하나 이 추출물에는 kanamycin에 대한 phosphorylation 효소는 존재하나 acetylation 효소는 존재하지 않은 것으로 확인되었다. 다른 kanamycin 생성균주인 S. kaηamyceticus NRRL 2535는 neomycin과 kanamycin에 강한 내성을 보였으나 이들 항생물질은 acetylation 시키거나 phosphorylation 시키는 효소는 확인되지 않았다. Gentamicin 또는 slsomicin을 생성하는 Micromonospora spp. 들은 gentamicin, kanamycin 그리고 tobramycin에 강한 내성을 보였으나 어떤 종류의 aminoglycoside 항생물질도 phosphorylaton 시키거나 acetylation 시키는 효소는 확인되지 않았다. Aminoglycoside 항생물질을 생성하는 Streptomycetes 들은 자신들이 생성하는 항생물질 뿐만 아니라 유사한 구조를 갖는 다른 항생물질에 대하여서도 특이한 내성메카니즘을 소유하고 있는 것으로 확인되었다. The cell free extracts of streptomycin producers, 7 strains of S. griseus and 1 strain of S. galbus inactivate streptomycin by phosphorylation, and the resistant mechanism against streptomycin by phosphorylation seems to be popular in streptomycin producing organisms. The cell free extracts of neomycin and paromomycin producers, S. fradiae and S. rimosus forma paromomycinus inactivate neomycin not only by phosphorylation, but also by acetylation. The cell free extract of a kanamycin producer. S. kanamyceticus (IFO 13414) inactivates neomycin by phosphorylation and/or by acetylation. However it does not inactivate kanamycin by phosphorylation but does by acetylation. The cell free extracts of the other kanamycin producer. S. kanamyceticus (NRRL 2535) does not show any inactivation of neomycin as well as kanamycin by phosphorylation or by acetylation, although it shows strong resisance against the antibiotics. The cell free extracts of Micromonospora spp.. producers of gentamicin or sisomicin which show resistance against gentamicin, kanamycin and tobramycin do not possess any phosphorylating or acetylating enzymes for the aminoglycoside antibiotics employed in this study. The Streptomycetes producing aminoglycoside antibiotics seem to have characteristic resistance mechanisms against the antibiotics produced by themselves and against other similar antibiotics.

Aminoglycoside 항생물질을 생성하는 Streptomycetes 와 Micromonospora spp 들의 Aminoglycoside 항생물질 변형효소에 관한 연구

구양모,최석례,김공환,임번삼,이새배 ( Yang Mo Goo,Seok Rye Choi,Kong Hwan Kim,Bun Sam Lim,Sae Bae Lee ) 생화학분자생물학회 1989 BMB Reports Vol.22 No.4

The cell free extracts of streptomycin producers, 7 strains of S. griseus and 1 strain of S. galbus inactivate streptomycin by phosphorylation, and the resistant mechanism against streptomycin by phosphorylation seems to be popular in streptomycin producing organisms. The cell free extracts of neomycin and paromomycin producers, S. fradiae and S. rimosus forma paromomycinus inactivate neomycin not only by phosphorylation, but also by acetylation. The cell free extract of a kanamycin producer. S. kanamyceticus (IFO 13414) inactivates neomycin by phosphorylation and/or by acetylation. However it does not inactivate kanamycin by phosphorylation but does by acetylation. The cell free extracts of the other kanamycin producer, S. kanamyceticus (NRRL 2535) does not show any inactivation of neomycin as well as kanamycin by phosphorylation or by acetylation, although it shows strong resisance against the antibiotics. The cell free extracts of Micromonospora spp.. producers of gentamicin or sisomicin which show resistance against gentamicin, kanamycin and tobramycin do not possess any phosphorylating or acetylating enzymes for the aminoglycoside antibiotics employed in this study. The Streptomycetes producing aminoglycoside antibiotics seem to have characteristic resistance mechanisms against the antibiotics produced by themselves and against other similar antibiotics.

Inhibition Studies on L-Phenylalanine Ammonia-lyase of Rhodosporidium toruloides

장지영,구양모,김공환,Chang, Ji-Young,Goo, Yang-Mo,Kim, Kong-Hwan 생화학분자생물학회 1988 한국생화학회지 Vol.21 No.4

Among the structural analogs of L-phenylalanine, trans-cinnamic acid, o-, m-, p-hydroxycinnamic acid, trans-cinnamamide and L-phenylglycine were found to be competitive inhibitors of L-phenylalanine ammonia-lyase of Rhodosporidium toruloides and their Ki values were 0.016, 0.023, 0.008, 0.022, 0.012 and 0.005 mM, respectively. Other compounds investigated hardly inhibited the enzyme. The active sites of the enzyme is thought to be composed of three portions that interact with the aromatic ring, the carboxyl or carboamide group and the ${\alpha}$, ${\beta}$-unsaturated bond. L-phenylalanine의 구조적인 유사체들 중에 trans-cinnamic acid, o-, m-, p-hydroxycinnamamide와 L-phenylglycine들이 Rhodosþoridium toruloides의 L-phenylalanine ammonia-lyase의 경쟁적인 반응억제제로 발견되었다. 그리고 그달의 Ki 값은 각각에 대하여 0.016, 0.023, 0.008, 0.022, 0.012 그리고 0.005 mM 이었다. 조사한 다른 많은 구조적인 유사체들은 그 효소의 반응속도를 거의 억제하지 않았다. 효소의 활성부위는 3부위로 구성되어 있는데 이들은 애로메틱고리와 산기 또는 아미드기 그리고 ${\alpha}-{\beta}-$ 불포화 결합과 상후 작용하는 것으로 생각된다.

Rhodosporidium toruloides 의 L - phenylalanine ammonia - Lyase 의 반응억제제에 관한 연구

장지영,구양모,김공환 ( Ji Young Chang,Yang Mo Goo ) 생화학분자생물학회 1988 BMB Reports Vol.21 No.4

Among the structural analogs of L-phenylalanine, trans-cinnamic acid, o-, m-, p-hydroxycinnamic acid, trans-cinnamamide and L-phenylglycine were found to be competitive inhibitors of L-phenylalanine ammonia-lyase of Rhodosporidium toruloides and their Ki values were 0.016, 0.023, 0.008, 0.022, 0.012 and 0.005 mM, respectively. Other compounds investigated hardly inhibited the enzyme. The active sites of the enzyme is thought to be composed of three portions that interact with the aromatic ring, the carboxyl or carboamide group and the α,β-unsaturated bond.

아미노페닐 아세토나이트릴의 정량 분석 : 가수분해 산물들 , 단백질 , 유화수소화합물들과 생체 시료들이 분석에 미치는 영향

이영복,구양모 ( Young Bok Lee,Yang Mo Goo ) 생화학분자생물학회 1986 BMB Reports Vol.19 No.4

Quantitative analysis of α-aminophenylacetonitrile was examined by analyzing cyanogen bromide formed by incubation of a sample in strong acidic solution with bromine. The reaction of the cyanogen bromide with pyridine followed by benzidine gave a substance showing a maximum absorption at 530 ㎚. The intensity of this absorption was linearly increased with concentration of α-aminophenylacetonitrile in the range of 0.01-0.2 mM. Addition of mercaptoethanol, or phenylglycine in the sample did not affect so much on colorization, but when albumin or phenylglycinamide was existing in the solution, the absorption at 530 ㎚ was found linearly increased with its concentration. Sulfur compounds, cysteine, methionine, glutathione and thioglycolic acid did not interfere in the analysis of α-aminophenylacetonitrile. Accuracy of the analytical methods for α-aminophenylacetonitrile in various biological samples were further examined.

Examination of Analytical Methods for Quantitative Analysis of $\alpha$-Aminophenylacetonitrile in the Presence of Its Hydrolyzed Products, Phenylglycine and Phenylglycinamide, and Proteins and Thiol Compounds, and in the Biological Samples

이영복,구양모,Lee, Young-Bok,Goo, Yang-Mo 생화학분자생물학회 1986 한국생화학회지 Vol.19 No.4

$\alpha$-Aminophenylacetonitrile의 농도를 브롬을 가한 강산과 함께 흔들어 주어 생성되는 cyanogen bromide를 분석하여 이루는 정량방법을 조사하였다. Cyanogen bromide는 pyridine과 함께 반응시킨 후에 benzidine을 처리하면 530 nm에서 최대의 흡광을 갖는 화합물이 얻어진다. 이 흡광은 0.01-0.2 mM의 농도 범위에서 $\alpha$-aminophenylacetonitrile의 농도에 대하여 비례적으로 증가하였다. 시료에 2-mercaptoethanol, 또는 phenylglycine을 가하여 주어도, 붉은 색의 생성에 큰 영향을 미치지 않았으나, 알부민이나 phenylglycinamide가 시료에 존재하는 경우 530 nm에서 흡광이 이들의 농도에 따라 직선적으로 증가하였다. 황화합물들인 cysteine, methionine, glutathione과 thioglycolic acid는 $\alpha$-aminophenylacetonitrile의 분석에 관여하지 아니하였다. 여러 형태의 생물학적인 시료내에 $\alpha$-aminophenylacetonitrile의 농도 정량에서 정밀도를 조사하여 보았다. Quantitative analysis of $\alpha$-aminophenylacetonitrile was examined by analyzing cyanogen bromide formed by incubation of a sample in strong acidic solution with bromine. The reaction of the cyanogen bromide with pyridine followed by benzidine gave a substance showing a maximum absorption at 530 nm. The intensity of this absorption was linearly increased with concentration of $\alpha$-aminophenylacetonitrile in the range of 0.01-0.2 mM. Addition of mercaptoethanol, or phenylglycine in the sample did not affect so much on colorization, but when albumin or phenylglycinamide was existing in the solution, the absorption at 530 nm was found linearly increased with its concentration. Sulfur compounds, cysteine, methionine, glutathione and thioglycolic acid did not interfere in the analysis of $\alpha$-aminophenylacetonitrile. Accuracy of the analytical methods for $\alpha$-aminophenylacetonitrile in various biological samples were further examined.

2-옥소카르바페남-3-카르보니트릴의 합성

백성인,이윤영,구양모,Baek, Seong In,Lee, Youn Young,Goo, Yang Mo 대한화학회 1995 대한화학회지 Vol.39 No.11