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Fate of Heavy Metals in Activated Sludge: Sorption of Heavy Metal ions by Nocardia amarae

  • Kim, Dong-wook
    • 한국환경과학회:학술대회논문집
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    • 한국환경과학회 1998년도 가을 학술발표회 프로그램
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    • pp.2-4
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    • 1998
  • Proliferation of Nocardia amarae cells in activated sludge has often been associated with the generation of nuisance foams. Despite intense research activities in recent years to examine the causes and control of Nocardia foaming in activated sludge, the foaming continued to persist throughout the activated sludge treatment plants in United States. In addition to causing various operational problems to treatment processes, the presence of Nocardia may have secondary effects on the fate of heavy metals that are not well known. For example, for treatment plants facing more stringent metal removal requirements, potential metal removal by Nocardia cells in foaming activated sludge would be a welcome secondary effect. In contrast, with new viosolid disposal regulations in place (Code o( Federal Regulation No. 503), higher concentration of metals in biosolids from foaming activated sludge could create management problems. The goal of this research was to investigate the metal sorption property of Nocardia amarae cells grown in batch reactors and in chemostat reactors. Specific surface area and metal sorption characteristics of N. amarae cells harvested at various growth stages were compared. Three metals examined in this study were copper, cadmium and nickel. Nocardia amarae strain (SRWTP isolate) used in this study was obtained from the University of California at Berkeley. The pure culture was grown in 4L batch reactor containing mineral salt medium with sodium acetate as the sole carbon source. In order to quantify the sorption of heavy metal ions to N amarae cell surfaces, cells from the batch reactor were harvested, washed, and suspended in 30mL centrifuge tubes. Metal sorption studies were conducted at pH 7.0 and ionlc strength of 10-2M. The sorption Isotherm showed that the cells harvested from the stationary and endogenous growth phase exhibited significantly higher metal sorption capacity than the cells from the exponential phase. The sequence of preferential uptake of metals by N. amarae cells was Cu>Cd>Ni. The specific surFace area of Nocardia cells was determined by a dye adsorption method. N.amarae cells growing at ewponential phase had significantly less specific surface area than that of stationary phase, indicating that the lower metal sorption capacity of Nocardia cells growing at exponential phase may be due to the lower specific surface area. The growth conditions of Nocardia cells in continuous culture affect their cell surface properties, thereby governing the adsorption capacity of heavy metal. The comparison of dye sorption isotherms for Nocardia cells growing at various growth rates revealed that the cell surface area increased with increasing sludge age, indicating that the cell surface area is highly dependent on the steady-state growth rate. The highest specific surface area of 199m21g was obtained from N.amarae cell harvested at 0.33 day-1 of growth rate. This result suggests that growth condition not only alters the structure of Nocardia cell wall but also affects the surface area, thus yielding more binding sites of metal removal. After reaching the steady-state condition at dilution rate, metal adsorption isotherms were used to determine the equilibrium distributions of metals between aqueous and Nocardia cell surfaces. The metal sorption capacity of Nocardia biomass harvested from 0.33 day-1 of growth rate was significantly higher than that of cells harvested from 0.5- and 1-day-1 operation, indicatng that N.amarae cells with a lower growth rate have higher sorpion capacity. This result was in close agreement with the trend observed from the batch study. To evaluate the effect of Nocardia cells on the metal binding capacity of activated sludge, specific surface area and metal sorption capacity of the mixture of Nocardia pure cultures and activated sludge biomass were determined by a series of batch experiments. The higher levels of Nocardia cells in the Nocardia-activated sludge samples resulted in the higher specific surface area, explaining the higher metal sorption sites by the mixed luquor samples containing greater amounts on Nocardia cells. The effect of Nocardia cells on the metal sorption capacity of activated sludge was evaluated by spiking an activated sludge sample with various amounts of pre culture Nocardia cells. The results of the Langmuir isotherm model fitted to the metal sorption by various mixtures of Nocardia and activated sludge indicated that the mixture containing higher Nocardia levels had higher metal adsorption capacity than the mixture containing lower Nocardia levels. At Nocardia levels above 100mg/g VSS, the metal sorption capacity of activate sludge increased proportionally with the amount of Noeardia cells present in the mixed liquor, indicating that the presence of Nocardia may increase the viosorption capacity of activated sludge.

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우리나라 어머니의 자녀 양육의 의미 - 거제지역을 대상으로 - (An Ethnography of Child-Rearing Experiences of Korean Mothers Living on Koje Island)

  • 이수연
    • 여성건강간호학회지
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    • 제7권4호
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    • pp.518-535
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    • 2001
  • Nursing practices should be based on the understanding of human beings. In order to understand human beings, it is important to study the lifestyles and thoughts of people in their natural environment. In this sense, the cultural aspects of a society need to be studied for a culture-bound nursing service. Child care, which is an important element of nursing, is also strongly influenced by the culture of a society. Therefore, a cultural study is necessary to understand the child-rearing practices of any society. The major purpose of this dissertation is to provide basic foundations for developing a culture-based theory for nursing intervention through studying traditional cultural elements of child care in Korean society. The study examined child-rearing practices in a small village on Koje Island in the southern part of Korea. It utilized ethnographic methodologies including participatory observations and in-depth interviews. The study participants were 9 Korean mothers living on Koje Island. The average age was 52. The data were collected between July in 1998 and December in 1999. The average number of interviews per person was 7-8, and the duration of each interview was approximately 2 hours. The data were analyzed using the Spradley Analytical Method. The following 9 major child-rearing aspects of mothers on Koje Island were discovered as a result of the study: 1. Firstly, mothers on Koje Island were mostly concerned about the "Old Birth Goddess' Curse", especially during their child's early years. This concern was evidenced by their careful behavior when their child was very young and by their praying to the Old Birth Goddess not to be jealous of their babies. 2. Secondly, they wished their children to live a different and better life than themselves. It was represented by their strong motivation toward their children's education as well as their expectation for their children's success. In traditional Korean culture, Korean people think that the rise and fall of the household depend on their offsprings. Therefore, Korean mothers wish their children attain to a higher level of social status through education. 3. Third, mothers are concerned about their children's righteousness. Mothers on Koje island expect their children to live with discretion, justice, strength, respect, harmony, and to do their best in life. 4. Next was an 'anticipation of their children's happy marriage'. The attributes of this category were an 'anxiety about their children's married life', and 'an expectation of a good spouse for their children'. Because Korean people believe that only a son can continue the bloodline of a family, especially Korean mothers have a great concern of the possibility of their daughters not having a son after marriage. Also they have different expectations toward their daughter-in-laws than son-in-laws. 5. Korean mothers also derived their satisfaction from their son. It was characterized by 'excessive affection toward their son', 'dependency on their son', and 'being afraid of their married daughter having a girl like themselves'. Korean society has been a patriarchy. Therefore, a son is beloved as someone who will take care of his old parents, be in charge of ancestral rites, and provide a daughter-in-law who can conceive a son. 6. The sixth category concerned 'the differences in their expectations for their children'. The attributes in this category were 'different expectations depending on their children's gender', 'different expectations depending on their children's ability', and a 'great sympathy toward children with low abilities'. Korean mothers expect their son to become better than their daughter. 7. The seventh category was related to their 'roles in child-caring practices'. Traditionally a child was raised in an extended family system in Korea So it was not the sole duty of a mother to bring up the child. Korean mothers used to receive much help rasing children from their in-laws, and family members. On the other hand, many children grew up by themselves, because their mothers were very busy taking care of housework. Furthermore, many children also grew up in poverty. 8. Mothers also had issues related to 'conflicts in child rearing'. They were characterized by 'lack of understanding', 'rudeness of children', and 'giving vent to one's anger'. 9. Finally, mothers regretted not doing their best in child-rearing practices. It was characterized by a 'bitter feeling of repentance', 'feeling irritated', and 'feeling of unsatisfaction'.

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Effects of Single Cell Protein Replacing Fish Meal in Diet on Growth Performance, Nutrient Digestibility and Intestinal Morphology in Weaned Pigs

  • Zhang, H.Y.;Piao, X.S.;Li, P.;Yi, J.Q.;Zhang, Q.;Li, Q.Y.;Liu, J.D.;Wang, G.Q.
    • Asian-Australasian Journal of Animal Sciences
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    • 제26권9호
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    • pp.1320-1328
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    • 2013
  • Three experiments were conducted to evaluate the ME value, standardized ileal digestibility (SID) of amino acids (AA) of fish meal, and the effects of single cell protein (Prosin and Protide) replacing fish meal in diet on growth performance, nutrient digestibility and intestinal morphology in weaned piglets. In Exp. 1, twenty-four barrows with initial BW of $30.8{\times}2.6kg$ were allotted to one of four dietary treatments. Diet 1 contained corn as the only energy source. The other three diets replaced 20% of the corn in diet 1 with one of the three protein feeds (fish meal, Prosin and Protide), and the DE and ME contents were determined by difference. In Exp. 2, eight barrows (initial BW of $25.6{\pm}3.2kg$) were fitted with ileal T-cannulas and allotted to a replicated $4{\times}4$ Latin square design. Three cornstarch-based diets were formulated using each of the protein feeds as the sole source of AA. A nitrogen-free diet was also formulated to measure endogenous losses of AA. In Exp. 3, one hundred and eighty piglets (initial BW of $7.95{\pm}1.59kg$) weaned at $28{\times}2d$ were blocked by weight and assigned to one of five treatments for a 28-d growth performance study, each treatment was fed to six pens with six pigs (three barrows and three gilts) per pen. The five treatments consisted of the control group (CON), which was a corn-soybean meal diet containing 5% fish meal, and the other four treatments, which replaced a set amount of fish meal with either Prosin (2.5% or 5%) or Protide (2.5% or 5%). The diets were formulated to provide same nutrient levels. The results showed that on a DM basis, both of the DE and ME contents were lower in Prosin and Protide than that of fish meal (p<0.05). The SID of CP and all essential AA were greater in fish meal than in Prosin and Protide (p<0.05). The pigs fed CON diet had greater weight gain and lower feed conversion rate (FCR) than pigs fed 5% Prosin and 5% Protide diets (p<0.05). The digestibility of CP was greater in pigs fed CON, 2.5% Prosin and 2.5% Protide diets than the pigs fed 5% Prosin and 5% Protide diets (p<0.05). Villus height in jejunum and ileum, and villus height to crypt depth ratio in the jejunum were higher (p<0.05) in pigs fed CON, 2.5% Prosin and 2.5% Protide diets compared with the 5% Prosin and 5% Protide diets. Pigs fed CON diet had greater villus height to crypt depth ratio in the ileum than the pigs fed 5% Prosin and 5% Protide diets (p<0.05). In conclusion, although Prosin and Protide contained lower ME content and SID of AA than fish meal, Prosin and Protide replacing 50% of fish meal in diet with identical nutrient levels could obtain similar performance, nutrient digestibility and intestinal morphology in weaned pigs.

방향족 화합물인 Aniline, benzoate, p-Hydroxybenzoate를 분해하는 Delftia sp. JK-2에서 분리된 Dioxygenases의 특성연구 (Characterization of different Dioxygenases isolated from Delftia sp. JK-2 capable of degrading Aromatic Compounds, Aniline, Benzoate, and p-Hydroxybenzoate)

  • 오계헌;황선영;천재우;강형일
    • KSBB Journal
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    • 제19권1호
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    • pp.50-56
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    • 2004
  • 본 연구의 목적은 방향족 화한물인 aniline, benzoate, p-hydroxybenzoate를 분해할 수 있는 Delftia sp. JK-2에서 이들 각 기질에서 배양시 다른 종류의 dioxygenases를 분리 정제하고, 정제된 dioxygenases의 특성을 조사하기 위하여 실시하기 위한 것이다. 기질로서 benzoate, aniline, 또는 p-hydroxybenzoate에 따라 분리된 dioxygenases는 각각 catechol 1,2-dioxygenase (C1 ,2O), catechol 2,3-dioxygenase(C2, 3O), 그리고 protocatechuate 4,5-dioxygenase (4,5-PCD)였다. 각 dioxygenases의 특성을 조사하기 위하여 먼저 benzoate, aniline 또는 p-hydroxybenzoate에서 배양한 Delftia sp. JK-2 세포를 초음파 분쇄기로 파쇄하여, ammonium sulfate precipitation, DEAE-sepharose, 그리고 Q-sepharose의 순서로 정제하여 농축하였다. 정제$.$농축된 dioxygenases의 특이 활성도를 보면 C1, 2O는 3.3 unit/mg, C2, 3O는 4.7unit/mg이고, 4,5-PCD는 2.0 unit/mg이다 C1, 2O와 C2, 3O의 기질 특이성 조사에서는 catechol과 4-methylcatechol에서 두 효소 모두 효소 활성이 나타났으며, C1. 2O에서는 3-methylcatechol에서 약간의 활성이 확인되었고, 4,5-PCD는 protocatechuate에서만 효소 활성을 보여주었다. C1l, 2O와 C2, 3O는 3$0^{\circ}C$와 pH 8.0에서 최적의 활성을 나타내는 것으로 조사되었으며, 4,5-PCD는 3$0^{\circ}C$와 pH 7.0에서 최적의 활성이 조사되었다. Delftia sp. JK-2에서 정제된 C1, 2O와 C2, 3O의 효소활성은 Ag$^{+}$, Hg$^{+}$, 그리고 Cu$^{2+}$에 의해 억제되는 것으로 나타났으며, 4,5-PCD의 경우에는 Ag$^{+}$, Hg$^{+}$, 그리고 Cu$^{2+}$ 뿐만 아니라 Fe$^{3+}$ 에 이해서도 효소 활성이 억제되는 것이 확인되었다. C1, 2O, C2, 3O, 4,5-PCD의 분자량은 SDS-PAGE에 의해 각각 60kDa, 35kDa, 62kDa로 측정되었다.

섬유 폐수 활성 슬러지에서 분리한 Comamonas testosteroni의 생물학적 페놀 분해 (Biodegradation of Phenol by Comamonas testosteroni DWB-1-8 Isolated from the Activated Sludge of Textile Wastewater)

  • 권해준;최두호;김미경;김동현;김영국;윤혁준;김종국
    • 생명과학회지
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    • 제30권2호
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    • pp.156-161
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    • 2020
  • 산업화 이후 다양한 화학물질의 생산과 이용은 우리 삶의 질을 높이는데 기여하였지만 그로 인한 대량의 폐기물 방출은 필연적이며 환경오염은 날이 갈수록 심각해지고 있다. 환경오염에 의한 화학물질의 노출은 인간의 건강과 생태계에 악영향을 주고 있다. 우리의 삶에 영향을 줄 수 있는 오염된 환경을 정화하는 것은 매우 중요한 문제이다. 광범위한 석유화학제품의 사용으로 인해 독성 난분해성 방향족화합물이 토양, 지하수, 폐수 등에서 빈번하게 검출되고 있다. 특히 합성수지, 합성섬유, 염료, 살충제, 방부제 등의 원료로 사용되는 페놀은 주요 오염물질이며, 호흡곤란, 두통, 구토, 돌연변이, 발암 등을 일으킬 수 있다. 본 연구에서는 페놀분해균주인 DWB-1-8을 섬유폐수에서 분리, 동정하였으며 16S rRNA유전자 염기서열분석결과 Comamonas testosteroni로 동정 되었다. 본 실험 균주의 최적 생장 및 최적페놀분해 배양 조건은 0.7% K2HPO4, 0.6% NaH2PO4, 0.1% NH4NO3, 0.015% MgSO4·7H2O, 0.001% FeSO4·7H2O, 초기 pH 7 및 30℃ 였으며, 다른 독성 화합물인 벤젠, 톨루엔 혹은 크실렌(BTX)을 유일탄소원으로 이용하여 생장할 수 있었다.

석유(탄화수소) 이용미생물에 관한 연구(제 1보) -효모세포에 의한 석유로부터 단백질 생성에 관하여- (Studies on the Petroleum hydrocarbon-utilizing Microorganisms(Part 1) -On the Production of Protein from the Yeast-cell-)

  • 이계호;신현경
    • Applied Biological Chemistry
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    • 제13권1호
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    • pp.43-50
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    • 1970
  • 석유탄화수소 이용미생물연구의 일환으로서 미생물에 의한 세포단백질생산의 목적으로 전국각지 135 개지역에서 주유소 세차장의 유침(油浸)토양을 비롯 석탄, 논밭, 하천토양등 242종을 수집하고 이시료로부터 석유탄화수소를 유일한 유기탄소원으로 이용하는 효모 468균주를 분리하였고 분리된 효모균주의 증식율(增殖率)을 screening 하여 우수균주를 선정하였다. 선정된 효모에 대하여 동정(同定) 및 배양 최적 조건의 검토 그리고 효모균체의 성분분석을 한 결과 다음과 같다. 1) 석유탄화수소 이용효모중 90.8%가 주유소및 세차장등 유침토양에서 나머지 10%가 석탄, 작토(作土) 하천토양에서 분리되였다. 2) 분리선정된 가장 우수한 효모균주는 주유소의 유침토양에서 분리됐고 Candida curvata HY-69-19로 동정되었다. 3) 분리선정된 Candida curvata HY-69-19는 optimum pH 5.0, optimum temperature $28^{\circ}C$, aerobic condition에서 균증식율이 컸다. 4) Candida curvata HY-66-19는 유기탄소원으로 petroleum fraction 중 비중이 0.8654이고 비점이 $268.9^{\circ}C$ 이상의 fraction인 heavy gas oil을 잘이용하며 무기질소원으로$(NH_2)_2CO$가 증식율과균체세포생산에 최적임을 알었다. 5) 분리선정된 이효모는 heavy gas oil 배지에서 lag phase 18시간, logarithmic growth phase 24시간${\sim}$42시간 사이며 이 때의 generation time은 3.8${\sim}4.5$시간이었다. 6) 분리선정된 이 효모는 heavy gas oil 및 $(NH_2)_2CO$ 배지로 54시간 pH 를 6시간마다 조정하면서 진탕배양하여 300mg/ml. H.G,0.의 건조균체를 생산하였다. 7) Candida curvata HY-69-19의 균체성분은 조단백질 40.25%, 조지방 14.81%,탄수화물 24.32% 회분 10.63%이었다.

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국내 분포 보리호위축바이러스(Barley Yellow Mosaic Virus) strain에 대한 저항성 유전자 반응 (Responses of Resistant Genes to Barley Yellow Mosaic Virus (BaYMV) Strains in Korea)

  • 박종철;노태환;박철수;강천식;강미형;이은숙;이준희;이정준;김태수
    • 식물병연구
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    • 제15권2호
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    • pp.72-76
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    • 2009
  • 국내 보리 재배지에 분포하는 BaYMV strain에 대한 BaYMV 저항성 유전자별 반응을 조사하였다. 지역별 바이러스 검정 결과 BaYMV가 전체 약 51%로 가장 높은 발생을 보였으며 BaMMV와의 혼합감염도 38.8%의 높은 비율을 나타내었다. BaMMV 단독 감염은 1.4%로 낮게 조사되었다. BaYMV 4개의 strain별 저항성 유전자의 반응을 조사한 결과 BaYMV-N type에 대해 검정한 결과 rym 1+5(Mokusekko-3), rym 3(Ea 52, Baitori), rym 5a(Solan)가 저항성을 보였다. -H strain은 rym 2(Mihori Hadaka 3), rym 3(Ea 52, Haganemugi, Baitori), rym 4m (Diana, Franka), rym 5a(Solan), rym 7(Hor 3365), rym 9 (Bulgarian 347), rym 12(Jochiwon Covered 2) 등 7종의 유전자가 저항성을 보여 다른 strain에 비해 효율적인 저항성이 많게 나타났다. -I strain의 경우 rym 1+5(Mokusekko-3), rym 3(Ea 52, Baitori), rym 5a(Hakei I-41, Solan) 등 3종의 유전자가 저항성을 보였다. -M starin에서는 rym 3(Chosen, Ea 52, Haganemugi, Baitori, Ishukushirazu), rym 4m(Diana), rym 5a(Hakei I-41) 등 3종의 유전자가 저항성을 보였다.

폐광지역에서 분리한 Benzoate 분해세균 Pseudomonas sp. NEQ-1에서 정제된 Catechol 1,2-Dioxygenase의 특성 (Characterization of Catechol l,2-Dioxygenase Purified from the Benzoate Degrading Bacterium, Pseudomonas sp. NFQ-l Isolated from Dead Coal Pit Areas)

  • 주정수;윤경하
    • 미생물학회지
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    • 제40권4호
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    • pp.275-281
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    • 2004
  • Quinoline (2,3-benzopyridine)을 유일한 탄소원과 질소원, 그리고 에너지원으로 이용하는Pseudomonas sp. NEQ-1을 실험 균주로 사용하였으며, 균주로부터 catechol 1,2-dioxygenase (C1,2O)를 유도하기 위하여 탄소원으로 benzoate를사 용하였다. C1,2O의 효소학적 특징을 조사하기 위하여 benzoate에서 배양한 Pseudomonas sp. NFQ-1을 초음파 분쇄기로 파쇄하고, ammonium sulfate침전과 gel permeation chromatography및 Source 15Q의 과정을 실시하여 C1,2O를 분리 및 정제하였다. 정제된 C1,2O의 특이활성(specific activity)은 14.21 unit/mg으로 나타났으며, SDS-PAGE에 의해 조사된 C1,2O의 분자량은 약 33 kDa이었다. Cl,2O는 catechol과 4-methylcatechol 및 3-methylcatechol에 대해서 효소활성을 나타내는 것으로 확인되었다. C1,2O의 Km은 38.54 ${\mu}M$로 측정되었고, Vmax는 $25.10\;{\mu}mol{\cdot}min^{-1}{\cdot}mg^{-1}$으로 나타났다. C1,2O는 $30^{\circ}C$와 pH 8.5에서 최적활성을 나타내는 것으로 조사되었으며, $Ag^+,\;Hg^+,\;Ca^{2+}$,그리고 $Cu^{2+}$는 C1,2O의 활성을 억제하였다. 분석되어진 N-말단 아미노산 서열은 ^1TVKISQSASIQKFFEEA^{17}$이었으며, Pseudomonas aeruginosa PA01과 $82\%$로 가장 높은 유사성을 보였고 Pseudomonas arvilla C-1와는 $71\%,$ Pseudomonas putida KT2440과는 $59\%,$ 그리고 Pseudomonas sp. CA10과는 $53\%$의 상동성이 각각 존재하는 것으로 확인하였다.

Burkholderia sp. D5에 의한 phenanthrene과 pyrene 분해 (Degradation of Phenanthrene and Pyrene by Burkholderia sp. D5)

  • 김태정;조경숙;류희욱
    • 미생물학회지
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    • 제39권4호
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    • pp.267-271
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    • 2003
  • Polycyclic aromatic hydrocarbons(PAHs)를 분해하는 Burkholderia sp. D5를 유류 오염 토양으로 분리하였고, PAHs의 분해특성을 조사하였다. 분리균주는 유일 탄소원으로 phenanthrene(Phe)을 이용하여 생장이 가능하였고, pyrene (Pyr)을 유일 탄소원으로 이용하여 생장하지는 못하였으나 yeast extract(YE)를 공기질로 첨가해 준 조건에서는 Pyr를 분해할 수 있었다. 분리 균주의 PAH 분해속도는 YE, peptone 및 glucose의 첨가에 의해 향상되었으며, 특히 YE 첨가 효과가 가장 우수하였다. 무기염배지(BSM)에 215 mg-Phe/L와 1 g-YE/L를 첨가한 조건에서 Burkholderia sp. D5의 비생장속도(0.28/h)와 Phe 분해속도(29.30 ${\mu}mol$/L/h)는 YE를 첨가하지 않은 조건에서 얻은 비생장속도(0.02/h)와 Phe 분해속도(12.00 ${\mu}mol$/L/h)의 각각 10배 및 2배였다. Phe를 기질로 공급한 경우 최대 비생장속도(${\mu}$_max)와 최대 Phe분해속도(V_max)는 각각 0.34 h-1 및 89 ${\mu}mol$/L/h 이었고. Pyr을 기질로 공급한 경우 ${\mu}$_max와 V_max는 각각 0.27 h-1 및 50 ${\mu}mol$/L/h이었다. Phe 혹은 Pyr 단독 기질하에서 분해속도와 비교 하였을 때(29.30 ${\mu}mol$-Phe L/h, 9.58 ${\mu}mol$-Pyr L/h), 두 기질의 혼합조건에서 Phe와 Pyr 분해속도는 각각 2.20 및 2.18 ${\mu}mol$ L/ h로 저하되었다. Burkholderia sp. D5 균주는 토양에서 Phe와 Pyr을 분해할 수 있었는데, Phe와 Pyr 분해속도는 각각 20.03 및 1.09 ${\mu}mol$ L/h 이었다.

Role of Citrate Synthase in Acetate Utilization and Protection from Stress-Induced Apoptosis

  • Lee, Yong-Joo;Kang, Hong-Yong;Maeng, Pil Jae
    • 한국미생물학회:학술대회논문집
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    • 한국미생물학회 2008년도 International Meeting of the Microbiological Society of Korea
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    • pp.39-41
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    • 2008
  • The yeast Saccharomyces cerevisiae has been shown to contain three isoforms of citrate synthase (CS). The mitochondrial CS, Cit1, catalyzes the first reaction of the TCA cycle, i.e., condensation of acetyl-CoA and oxaloacetate to form citrate [1]. The peroxisomal CS, Cit2, participates in the glyoxylate cycle [2]. The third CS is a minor mitochondrial isofunctional enzyme, Cit3, and related to glycerol metabolism. However, the level of its intracellular activity is low and insufficient for metabolic needs of cells [3]. It has been reported that ${\Delta}cit1$ strain is not able to grow with acetate as a sole carbon source on either rich or minimal medium and that it shows a lag in attaining parental growth rates on nonfermentable carbon sources [2, 4, 5]. Cells of ${\Delta}cit2$, on the other hand, have similar growth phenotype as wild-type on various carbon sources. Thus, the biochemical basis of carbon metabolism in the yeast cells with deletion of CIT1 or CIT2 gene has not been clearly addressed yet. In the present study, we focused our efforts on understanding the function of Cit2 in utilizing $C_2$ carbon sources and then found that ${\Delta}cit1$ cells can grow on minimal medium containing $C_2$ carbon sources, such as acetate. We also analyzed that the characteristics of mutant strains defective in each of the genes encoding the enzymes involved in TCA and glyoxylate cycles and membrane carriers for metabolite transport. Our results suggest that citrate produced by peroxisomal CS can be utilized via glyoxylate cycle, and moreover that the glyoxylate cycle by itself functions as a fully competent metabolic pathway for acetate utilization in S. cerevisiae. We also studied the relationship between Cit1 and apoptosis in S. cerevisiae [6]. In multicellular organisms, apoptosis is a highly regulated process of cell death that allows a cell to self-degrade in order for the body to eliminate potentially threatening or undesired cells, and thus is a crucial event for common defense mechanisms and in development [7]. The process of cellular suicide is also present in unicellular organisms such as yeast Saccharomyces cerevisiae [8]. When unicellular organisms are exposed to harsh conditions, apoptosis may serve as a defense mechanism for the preservation of cell populations through the sacrifice of some members of a population to promote the survival of others [9]. Apoptosis in S. cerevisiae shows some typical features of mammalian apoptosis such as flipping of phosphatidylserine, membrane blebbing, chromatin condensation and margination, and DNA cleavage [10]. Yeast cells with ${\Delta}cit1$ deletion showed a temperature-sensitive growth phenotype, and displayed a rapid loss in viability associated with typical apoptotic hallmarks, i.e., ROS accumulation, nuclear fragmentation, DNA breakage, and phosphatidylserine translocation, when exposed to heat stress. Upon long-term cultivation, ${\Delta}cit1$ cells showed increased potentials for both aging-induced apoptosis and adaptive regrowth. Activation of the metacaspase Yca1 was detected during heat- or aging-induced apoptosis in ${\Delta}cit1$ cells, and accordingly, deletion of YCA1 suppressed the apoptotic phenotype caused by ${\Delta}cit1$ mutation. Cells with ${\Delta}cit1$ deletion showed higher tendency toward glutathione (GSH) depletion and subsequent ROS accumulation than the wild-type, which was rescued by exogenous GSH, glutamate, or glutathione disulfide (GSSG). Beside Cit1, other enzymes of TCA cycle and glutamate dehydrogenases (GDHs) were found to be involved in stress-induced apoptosis. Deletion of the genes encoding the TCA cycle enzymes and one of the three GDHs, Gdh3, caused increased sensitivity to heat stress. These results lead us to conclude that GSH deficiency in ${\Delta}cit1$ cells is caused by an insufficient supply of glutamate necessary for biosynthesis of GSH rather than the depletion of reducing power required for reduction of GSSG to GSH.

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