Purification and Characteristic Properties of Ribonuclease H from Sea Urchin, Hemicentrotus pulcherrimus

류병호,이병호,박형숙,하미숙,김동석,Ryu, Beung-Ho,Lee, Beung-Ho,Park, Hyeong-Suck,Ha, Mi-Suck,Kim, Dong-Seuk 생화학분자생물학회 1986 한국생화학회지 Vol.19 No.3

말똥 성게를 인공적으로 수정시킨 후, 칼럼 크로마토그래피법으로 분리 정제하여 리보뉴크레아제 $H_1$과 리보뉴크레아제 $H_2$를 얻었다. 리보뉴크레아제 $H_1$의 분자량은 약 68,000이었고, 리보뉴크레아제 $H_2$의 분자량은 약 44,000 이었다. 리보뉴크레아제 $H_1$는 약알카리성에서, 리보뉴크레아제 $H_2$는 알카리성에서 활성이 높았다. 이들 효소는 모두 2가이온인 마그네슘이온 ($Mg^{2+}$), 망간이온($Mn^{2+}$)과 그리고 염의 일정농도에서 활성을 나타내고 있으며, N-에칠 말례이마이드에 감수성이 높았다. 그리고, 리보뉴크레아제 $H_1$과 $H_2$는 단일가닥과 이중가닥의 활성은 없었으나, RNA-DNA hydrid는 급히 저하 시켰다. From the sea-urchin, Hemicentrotus pulcherrimus, we have purified by two column chromatographic steps two ribonuclease H (RNase H) activities of RNase $H_1$ and RNase $H_2$ in homogeneous form. RNase $H_1$ was molecular weight around 68,000 and RNase $H_2$ was around 44,000 by Sephadex G-75 gel filteration and SDS-polyacrylamid gel electrophoresis. RNase $H_1$ was optimal active as well as slightly alkaline pH and RNase $H_2$ was as alkaline pH. These enzymes shown to be a function of the divalent metal ion $Mg^{2+}$, $Mn^{2+}$ and salt employed as activator, also were sensitive -SH reagent N-ethylmaleimide. RNase $H_1$ and RNase $H_2$ were shown no activity with single or double stranded RNA but rapidly hydrolysis the RNA-DNA hybrid.

카드뮴 축적 변이주인 Pseudomonas maltophilia H - 8M의 특성

류병호(Beung-Ho Ryu),노명훈(Myung-Hoon Rho),정수자(Su-Ja Jung),배기철(Ki-ChuI Bae) 한국식품영양과학회 1992 한국식품영양과학회지 Vol.21 No.1

본 연구는 cadmium의 축적균주인 Pseudomonas maltophilia H-8을 MNNG로 처리하여 cadmium축적능이 가장 우수한 변이균주인 Pseudomonas maltophilia H-8M을 얻었고 그 특성을 조사하였다. 변이주 Pseudomonas maltophilia H-8M은 카드뮴농도 1,000ppm에서 36시간 배양했을 시 대조구에 비해 약 80%정도의 성장율을 나타내었다. 카드뮴 이외의 중금속의 첨가에 따른 생육상은 Pb^(2+), Zn^(2+), Hg^(2+), Cu^(2+), Cr^(2+), Co^(2+)등에는 별다른 성장 억제는 없었으나 Sn^(2+)에서는 생육억제가나타났다. 또한 50ppm의 카드뮴을 배지에 첨가 하였을 때 균체내 카드뮴 축적율은 52.92%로 가장 높게 나타났으며 축적부위는 세포벽에 80.3%의 축적율을 나타내었다. This study was carried out to investigate the characteristics of a mutant, Pseudomonas maltophilia Ho8M selected with the treatment of Pseudomonas maltophilia H-8 by N-methyl-N-nitro-N-nitrosoguanidine (MNNG). This mutant showed highest ability of cadmium accumulation. The growth rate of Pseudomonas maltophilia H-8M showed about 80% in 1000ppm Cd containing medium when compare with control for 36h at 30℃. Pseudomonas maltophilia H-8M not inhibited on the growth in addition of various heavy metal such as Hg^(2+), Zn^(2+), Pb^(2+), Cu^(2+), Cr^(2+) and Co^(2+), but inhibited in Sn^(2+) containing medium, respectively. Pseudomonas maltophilia H-8M was accumulated the highest cadmium level of 62.3% on whole cell in the medium containing 50 ppm and 80% of accumulated cadmium was distributed in the cell wall.

카페인 분해균주의 분리 및 특성

류병호,주신회,Ryu, Beung-Ho,Ju, Sin-Hae 한국식품과학회 1992 한국식품과학회지 Vol.24 No.3

Caffeine을 기질로 이용할 수 있는 균을 토양과 폐수에서 7종을 분리하고 그중 카페인 분해능이 가장 좋은 KS-5를 동정하여 Pseudomonas putida KS-5라고 명명하였다. P. putida KS-5의 생장 최적조건은 온도 $30^{\circ}C$, pH 7.0이었고 카페인 농도는 1.0%였다. P. putida KS-5는 plasmid DNA가 존재하였으며, 카페인 분해유전자가 plasmid에 존재할 수 있음을 알 수 있었다. 한편 이들 분해유전자의 유전적 특징을 규명하기 위한 curing test와 transformation에 필요한 marker를 찾아본 결과 각종 항생물질에 대하여 내성이 있었다. Several bacterial strains capable of degrading caffeine were isolated and studied for their biodegradation ability of the caffeine and some biochemical characteristics. The isolate KS-5 was identified as Pseudomonas putida and was designated as the P. putida KS-5. The optimum conditions were at $30^{\circ}C$, pH 7.0 and 1.0% caffeine. Agarose gel electrophoresis and curing experiment were found that the gene for caffeine degradation was encoded on the plasmid in P. putida KS-5 and that this strain was resistant to several antibiotics.

Aspergillus oryzae A-5로부터 Low Density Lipoprotein(LDL)의 산화에 대한 항산화 효과

류병호,김동석,조경자,이홍수,진성현,Ryu, Beung-Ho,Kim, Dong-Suck,Cho, Kyung-Ja,Lee, Hong-Su,Jin, Sung-Hyun 한국생명과학회 1997 생명과학회지 Vol.7 No.4

Antioxidative activity of fraction extracted from cultivation of Aspergillus sp. A-5 against oxidation of human low density lipoprotein(LDL) was investigated. Fractions of Aspergillus sp. A-5 cultivation was sucessively purified with ethyl acetate and silica gel column chromatography. The concentration of fraction 4 inhibited Cu$^{2+}$-induced oxidation of LDL almost completely. Band 3 isolated by the further purification of fraction 4 was higher than that of same concentration of $\alpha$-tocopherol, BHA and BHT. The elcetrophoretic mobility of oxidized LDL by addition of Band 3 was faster than that of native LDL, but slower than that of oxidzed LDL. It is concluded that fraction of Aspergillus cultivation contained antioxidants with the capacity to inhibit oxidative modification of LDL.

엉겅퀴에서 분리정제한 silybin의 Human Low Density Lipoprotein 수식에 대한 항산화 효과

류병호,김희숙,이백천,이홍수,Ryu, Beung-Ho,Kim, Hee-Suck,Lee, Baek-Cheon,Lee, Hong-Su 한국생명과학회 1998 생명과학회지 Vol.8 No.4

This study was initiated to antioxidant activity of silybin on oxidation of human low density lipoproteins(LDL). Siltbin was extracted from Silybum marianum by the combination of fractionation and it was futher purified by silica gel column chromatography, and isolated active substances were identified silybin by IR, NMR and GC-MS. siltbin inhibited the ozidation of human low density lipoprotein(LDL) mediated by 5$^{\mu}$m CU $^{2+}$ ion in a dose dependent manner. LDL oxidation by congugated dines formation was completely inhibited by silybin at a concentration of 5$^{\mu}$M. The results provide a possibility that silybin might protect LDL against oxidation in atherosclerotic lessions.

Antioxidative activity of flavonoid rich extract of Oenothers odorate Jacquin on oxidation of low density lipoprotein

류병호,김희숙,조경자,Ryu, Beung-Ho,Kim, Hee-Sook,Cho, Kyung-Ja Korean Society of Life Science 2002 생명과학회지 Vol.12 No.3

There is growing interest ill understanding the role and mechanisms of flavonoid as antioxidant on LDL. The antioxidative activity of flavonoid typically present in Oenothers odorate Jacquin was investigated in vitro using a human LDL oxidation assay. In present work, LDL was incubated with increasing concentrations of extracts of extracts of Oenothers odorate Jacquin and LDL oxidation was started by adding CuSO$_4$to the media. Substances in leaves extracts of Oenothers odorate Jacquin are capable of inhibiting the initiation and the propagation of LDL oxidation. They inhibit LDL oxidation, monitored by thiobarbituric acid-reactive substances(TBARS), as well as modification as shown through direct measurement of electrophoretic mobility, diene conjugates. Inhibition is a dose dependent effect that becomes already apparent at concentration of extracts as low as 40$\mu\textrm{g}$/mL. Inhibition is almost complete at 80$\mu\textrm{g}$/mL. Extracts of Oenothers odorate Jacquin were more potent antioxidative activity than either ascorbic acid and dl-a -tocopherol. 달맞이꽃 추출물로써 항산화활성을 알아보기 위하여 사람 low densty lipoprotein(LDL)에 대하여 실험하였다. 사람 LDL에 CuSO$_4$를 첨가한 후 배양하면서 달맞이꽃 추출을 일정 농도씩 증가시키면서 첨가하여 배양하였다. 달맞이꽃 추출물로서 LDL의 산화에 대한 개시와 연쇄 반응에서 thiobarbituric acid reactivy substances (TBARS)와 전기 영동 및 공액 2중 결합으로 측정하였다. 달맞이꽃 추출물을 LDL에 첨가한 후 macrophage J774로써 배양한 결과 TBARS값은 용량 의존형의 항산화 활성을 나타내었으며 전기 영동 이동상에서도 대조구에 비하여 각 시험구에서 LDL의 산화를 억제하였다. 또 공액 2중 결합에 대한 실험에서 용량 의존형산화 억제 효과를 나타내었다. 달맞이꽃 추출물의 항산화 활성은 40 $\mu\textrm{g}$/ml의 농도에서도 항산화 효과가 있었으나 80$\mu\textrm{g}$/mL의 농도에서는 거의 완전한 억제효과가 있었다. 그리고 비타민 C 및 $\alpha$-dl-tocopherol과 비교한 결과 달맞이들 추출물이 약간 높은 항산화 활성을 나타내었다.

Characteristics of Bluish Purple Pigment Produced by Streptomyces californicus KS - 89

류병호(Beung-Ho Ryu),지영애(Young-Eh Chi),이병호(Byeong-Ho Lee),박법규(Bub-Gyu Park),박우열(Woo-Yeol Park) 한국식품영양과학회 1990 한국식품영양과학회지 Vol.19 No.3

천연식용 색소의 개발을 위한 하나의 방안으로 Streptomyces californicus KS - 89에 의하여 생산되는 청자색 색소의 물리 화학적 특성을 조사하였다. 청자색 색소를 실리카겔의 칼럼 크로마토그래피에 의하여 분리하였고, C.I.E. chromatic diagram에 의하면 청자색에 속하며 UV 최대 흡수대는 575㎚이였다. 이 색소의 수용액에서의 색조는 pH 5~8 범위내에서는 안정하였고, 자외선에 의하여 영향을 받지 않으나, 때때로 열에 의하여 색상이 약간 흐려진다. 한편 금속염에는 거의 안정하였고, 특히 이 색소는 돌연변이원성과 항암 효과는 없었으며, 또 항생 물질능도 없었음이 밝혀졌다. Aqueous solution pigment produced by Streptomyces californicus KS-89 showed a vivid bluish purple pigment and purified by silica gel column chromatography. The pigment indicated a deep purple color zone by the C.I.E. chromatic diagram, and showed UV absorption maxima at 575㎚. The color intensity in aqueous solution was fairly stable in the ranges of pH 5~8 and was not affected by UV light, however, sometimes it had faded slightly by the heat. It was possible to prevent significantly by the addition of metal salt. Especially, this pigment has no mutagenicity and antitumor activity and it appears to be devoid of antibiotic activity.

Semicontinuous Production of Blue Pigment from Gardenia Fruit by Immobilized Cells of Bacillus subtilis KS - 380 Using Air Bubble Column Reactor

류병호(Beung-Ho Ryu),조경자(Kyung-Ja Cho) 한국식품영양과학회 1990 한국식품영양과학회지 Vol.19 No.5

치자로 부터 청색 색소를 생산하기 위하여 β-glucosidase를 분비하는 Bacillus subtilis KS-380을 고정화시켜 air bubble column reactor를 이용하여 free cells와 고정화 세포로서 반연속식 발효를 실시하였다. 청색 색소의 생산은 Bacillus subtilis KS-380 균체를 3.5% 알긴산 소다로 고정화시키고 gel bead는 2㎜크기로서 0.2㎖/min로 aeration 하였을 때 생산성이 가장 우수하였다. Air bubble column을 이용하여 최적 생산 조건에서 매회 24시간 간격으로 발효시 마다 새로운 생산 배지를 공급하면서 4회 반연속적으로 생산한 결과 free cells 보다는 매우 높은 생산성을 나타내었다. The semicontinuous production of blue pigment from gardenia fruit by immobilized cells of Bacillus subtilits KS-380 which excreted β-glucosidase was investigated in comparison with free cells. The blue pigment produced highest productivity under the conditions of aeration of 0.2㎖/min and 2㎜ diameter of gel beads by using 3.5% sodium alginate. Semicontinuous production by immobilized cells showed the highest productivity with replacement of fresh production medium in every 24hr for fourth fermentation cycle following the conditions of blue pigment productivity.

Monascus pilosus M - 15에 의한 콜레스테롤 합성저해제 Monacolin의 생산

류병호(Beung-Ho Ryu),안문규(Moon-Kyu Ahn),박종옥(Jeung-Ok Park) 한국식품영양과학회 1995 한국식품영양과학회지 Vol.24 No.1

본 연구는 콜레스테롤 합성저해제인 monacolin을 생산하기 위하여 Monascus sp.에서 생산균주를 선발하고 생산조건을 검토하였다. Monascus sp. 중에서 Mon-ascus pilosus M-15에 의하여 생산되는 monacolin의 생산조건은 최적 생산 배지로는 5% glycerin, 2% glucose, 0.5% corn steep liquor, 0.5% NaNO₃, 0.2% MgSO₄ㆍ7H₂O이였으며, 최적 pH는 6.0이였고, 25℃에서 10~20일간 배양하였을 때 monacolin을 추출하여 UV.IR. 및 GC/MS로 확인한 결과 표준품인 monacolin과 거의 비슷한 경향을 나타내었다. The monacolin retard in vivo synthesis of cholestrol. This study was conducted to select the best strain of Monascus sp. for monacolin production among 21 strains. In addition, research on suitable condition for mass production was conducted. Monascus pilosus M-15 was selected as the best strain for production of monacolin. The appropriate substrate for the monacolin production seem to be as follow : glycerin 5%, glucose 2%, corn steep liquor 0.5%, sodium nitrate 0.5% and length of fermentation was at 25℃ for 10~12days, respectively. The monacolin obtained from this organism was compared with standard monacolin using UV, IR, and GC/MS system and it was very close to the standard monacolin.

Aspergillus awamori, B-2의 균사체 고정화 의한 Melanoidin 탈색

류병호(Beung-Ho Ryu),김혜성(Hye-Sung Kim),하미숙(Mi-Suk Ha),정종순(Jong-Sun Jung),빈재훈(Jae-Hun Bin),이영숙(Young-Sook Lee),정수자(Soo-Ja Chung) 한국식품영양과학회 1989 한국식품영양과학회지 Vol.18 No.1

곰팡중에서 melanoidin의 탈색력이 있는 Aspergillus awamori, B-2을 선별하여 탈색력을 실험하였다. Aspergillus awamori, B-2는 3.0% glucose, 0.5% yeast extract, 0.1% KH₂PO₄ 및 0.05% MgSO₄ㆍ7H₂O을 함유하는 melanoidin 용액(pH 7.0) 에서 5일 동안 37℃에서 배양했을 때 가장 높은 탈색력을 나타내었다. 탈색력을 효과적으로 하기 위하여 Ca - alginate로 고정화하여 최적조건에서 계속적으로 탈색하였을대 mycelia 단독보다는 더욱 효과적이었다. Melanoidin 탈색력은 최적 조건에서 10일 동안 반응하였을때 약 70%을 나타내었다. 고정화한 mycelia의 재사용에 의한 계속적인 탈색을 시도하였던바 15일 동안 연속적으로 반응시켜도 약 60-70%의 탈색력을 계속 유지하였다. Aspergillus awamori, B-2 which has a high ability to decolorize melanoidin was selected among various fungi.<br/> Aspergillus awamori, B-2 showed the highest decolorization activity when it was cultivated in a melanoidin medium containing 3.0% glucose, 0.5% yeast extract, 0.1% KH₂PO₄and 0.05 % MgSO₄ㆍ7H₂O at an initial pH 7.0 at 37℃ for 5 days.<br/> Mycelia immobilized system with Ca-alginate was more effective on melanoidin decolorization activity showed approximately 70% in 10 days under the optimal conditions.<br/> Continuous decolorzation of melanoidin using reuse of immobilized mycelia showed an almost constant decolorization of abort 60-70 % for 15 days.