연속배양에서 고핵산 효모 변이주의 배양생리학적 특성

남수완 동의대학교 산업기술개발연구소 2005 産業技術硏究誌 Vol.19 No.-

In order to investigate the physiological characteristics for RNA accumulation, continuous cultures of S. cerevisiae MTY62 under the carbon-limitation or carbon and phosphate-limitation were performed. In the carbon-limited chemostat, the higher RNA concentration of 550 g-DCW/L and the RNA content of 144 mg-RNA/g-DCW cell was observed at the dilution rate of 0.30 hr^(-1). In the carbon- and phosphate-limited chemostat, the culture condition with the dilution rate of 0.25 hr^(-1) gave the maximal RNA content of 209 mg-RNA/g-DCW. The accumulation rate of RNA (mg-RNA/L·hr) was decreased at higher dilution rate in the carbon and phosphate-limited chemostat.

효모의 구성적 Promoter들에 의한 Inulinase 유전자의 발현

남수완,김연희 동의대학교 기초과학연구소 2000 基礎科學硏究論文集 Vol.10 No.1

To express constitutively the inulinase gene (INU1) of Kluyveromyces marxianus in Saccharomyces, cerevisiae, three yeast promoters such as GAPDH, ADH1 and ENO1 were connected upstream of INU1. The resulting plasmids, pYIGP, pADH1-INU, and pENO-INU were introduced to S. cerevisiae SEY2102 host strain, respectively, and then each transformants were selected by staining of colonies on sucrose-agar plate. When the yeast transformants were cultivated on 2% dextrose media, the total expression levels of inulinase reached to 1.11 unit/mL, 0.88 unit/mL, and 0.69 unit/mL for respective GAPDH, ADH1, and ENO1 promoter systems. On 4% dextrose media, however, the inulinase activities were observed at 2.00 unit/mL for pYIGP, 0.71 unit/mL for pADH1-INU, and 1.40 unit/mL for pENO-INU. This result indicates that the constitutive expression of INU1 was significantly affected by the initial concentration of dextrose and the promoter strength was in the order GAPDH, ENO1, and ADH1 promoter at high dextrose concentration. Taking into account the plasmid stability, however, it is suggested that the ENO1 promoter system is more suitable for the INU1 expression on high dextrose medium or in the fed-batch cultivation accumulating ethanol at high level.

재조합 Saccharomyces cerevisiae에서 Inulinase와 Invertase의 발현과 분비에 미치는 배양조건의 영향

남수완,신동하,김연희 동의대학교 기초과학연구소 1998 基礎科學硏究論文集 Vol.8 No.1

The effects of medium pH and culture temperature on the expression and secretion and secretion of inulinase and invertase were investigated with recombinant Saccharomyces cerevisiae cells. These cells were obtained by transformation of 2μ-based plasmids pYI10 and pYS10 which contain Kluyveromyces marxianus inulinase gene (INU1A) and S. cerevisiae invertase gene (SUC2), respectively, in the downstream of GAL1 promoter. The expression level and localization of inulinase and invertase were not affected significantly by the initial medium pH: secretion efficiencies of inulinase and invertase into the medium were about 90% and 60%, respectively, in the pH ranges of 4.0 to 6.5. However, the expression and secretion of both enzymes were strongly dependent on the culture temperature. The highest expression (7.7 units/mL) and secretion (6.7 units/mL) of inulinase were observed at 28℃ and 30℃. As a consequence of decreased localization of inulinase in the periplasmic space, the secretion efficiency increased from 68% at 20℃ to 95% at 35℃. The total expression level and secretion efficiency of invertase increased from 19 units/mL and 55% at 20℃ to 25 units/mL and 68% at 35℃, respectively. Irrespective of the culture temperature, the invertase activity in the cellular fraction (periplasmic space and cytoplasmic fractions) was kept constant at around 33∼45%.

Saccharomyces cerevisiae에서 Trichoderma Endoglucanase의 발현과 분비

남수완,김병우,신동하,김재범,신지원,정대균,정춘수 동의대학교 기초과학연구소 1999 基礎科學硏究論文集 Vol.9 No.1

The endoglucanase gene, egl6, of Trichoderma sp. was connected with the yeast ADH1 promoter, and the resultant plasmid, pVT-C4, was introduced into three S. cerevisiae host strains (YNN27, 2805, and SEY2102). Among each 80 transformants, the cell growth and expression level of endoglucanase were compared in test-tube cultivation, and three respective transformants for each host cells showing the highest expression level and cell growth were selected. When three recombinant yeast cells were batchwise cultivated for 48 hr in flask, the total activities of endoglucanase expressed were about 1140 unit/1 with 2805/pVT-C4, 1020 unit/l with SEY2102/pVT-C4, and 590 unit/l with (YNN27/pVT-C4. Irrespective of host strain, about 80% of the expressed endoglucanase was detected in the extracellular medium. In addition, it was also found that the recombinant enzyme was secreted into the culture medium as two major forms of lightly and heavily glycosylated proteins.

Escherichia coli 패혈증 환자에 합병된 대칭적 하지 말단 괴사증 1예

남해성,유진홍,권순석,민준기,조현선,박민경,심병주,남유정,이지인,김진수,길욱현,조근종,신완식 대한감염학회 2005 감염과 화학요법 Vol.37 No.6

We have encountered a rare case of symmetrical peripheral gangrene complicating Escherichia coli sepsis in a 47-years-old male. He was successfully treated with antibiotics, anticoagulants, and vasodilator. To our knowledge, this is the first report on symmetrical peripheral gangrene complicating E. coli sepsis in Korea.

Saccharomyces cerevisiae에서 Clostridium thermocellum Endoglucanase 유전자의 구성적 발현

남수완,정대균,정봉현 동의대학교 기초과학연구소 1998 基礎科學硏究論文集 Vol.8 No.1

To develop an effective and powerful yeast probiotics, Saccharomyces cerevisiae strains producing cellulolytic enzymes were genetically engineered. We constructed two plasmids in which the endoglucanase A gene, celA, of Clostridium thermocellum was connected in frame with ADH1 or GAPDH promoter. These plasmids were transformed into various S. cerevisiae host strains, and then the cell growth, expression level and plasmid stability between the transformed yeast cells were examined in the flask culture. The difference in genetic background of host strains did affect significantly the cell growth and expression level of endoglucanase. Based on the higher levels of cell growth(5.4∼5.9 g-DCW/L) and plasmid stability(79∼85%), three host cells(YNN27, 2805 and SEY2102) and ADH1 promoter were selected as optimal host-vector systems for the constitutive expression of celA. The recombinant yeast produced about 200 unit/L of endoglucanase as a growth-associated manner in the batch fermentation, due to the increased cell growth of 11∼13 g-DCW/L. In addition, 25∼38% and 53∼64% of the endoglucanase activity were detected in the culture medium and periplasmic space, respectively. These results indicate that the signal sequence of celA functioned well in S. cerevisiae cells and the recombinant yeast cells can be employed for the production of probiotics.

한약과 양약의 약물상호작용에 관한 연구(제1보) : Nicardipine과 청혈단의 상호작용 Effects of CHUNGHYULDAN on the Pharmacokinetics and Pharmacodydnamics of Nicardipine in Rats

김남재,진성민,최혁재,주수만,송보완,김종우 한국병원약사회 2002 병원약사회지 Vol.19 No.4

The influence of single and repeated oral pretreatment of CHUNG-HYUL-DAN(CHD). a widely used Korean traditional herbal medicines prescribed for hypertension and hyperlipidemia, on the pharmacokinetics and pharmacodynamics of a calcium channel blocker, nicardipine after oral administration of nicardipine was investigated in rats. The single pretreatment of CHD significantly increased area under the plasma concentration-time curve(AUC_(0-8h), but did not influence the time to reach the maximum peak plasma concentration(T_(max)), the maximum peak plasma concentration(C_(max)), terminal elimination half-life(t_(1/2) or the elimination rate constant(λ) for nicardipine. And, C_(max) and AUC_(0-8h) for nicardipine were significantly increased by with 1-week repeated pretreatment with CHD, but the other parameters were not significantly different from those of control. Also, the extent of the lowering effect of nicardipine on blood pressure in rats was significantly increased after oral administration of nicardipine with single and repeated oral pretreatment with CHD. These results indicated that single and 1-week repeated oral pretreatment with CHD should inhibit the metabolism of nicardipine in rats.

컨테이너 터미널 ERP 시스템 구축 방안

이상완,최형림,박남규,김현수,박병주,유동호 한국로지스틱스학회 2002 로지스틱스연구 Vol.10 No.1

급속한 세계화로 터미널간의 경쟁이 치열해지면서 기존 시설의 효율적인 운영 및 관리가 터미널의 경쟁력과 생산성을 향상시키는 중요한 요소로 부각되고 있다. 하지만 우리나라 컨테이너 터미널은 양적 확장에 치중한 반면 관리와 운영의 효율성에 대해서는 다소 소홀히 해 왔다. 그리고 대부분의 컨테이너 터미널은 자사 운영시스템을 저문 업체에 의뢰하여 개발함으로써 컨테이너 터미널마다 상이한 시스템이 구축되어 시스템의 유지 보수비용이 많이 발생하고 있다. 최근 터미널 개발과 운영 자체가 사업화 되고 있기에 향후 터미널 운영시스템에 대한 수요는 증가할 것이다. 그래서 터미널마다 다른 운영시스템을 표준화하고 국제화한 ERP 형태의 통합 운영 시스템의 개발이 요구된다 하겠다. 따라서 본 연구에서는 주문형 방식에 의존하던 컨테이너 터미널 운영시스템의 고비용, 비 통합화, 개발기간의 장기화 등의 단점을 극복하기 위해 컨테이너 터미널 ERP 시스템이란 새로운 개념을 제안한다. 이 컨테이너 터미널 ERP 시스템은 통합된 운영시스템 구축, 개발기간의 단출, 개발비용의 절감, 쉬운 업그레이드를 가능하게 해 컨테이너 터미널의 글로벌 사업에 전략적 도구로써 역할을 하게 될 것으로 기대된다. The major characteristics of ERP(Enterprise Resource Planning)are enterprise-wide system that covers all the business functions and information resources, integrated database, built-in best industry practice, packaged software and open architecture. ERP enables reduction of system development time, flexibility, standardization of workflow and effective business planning capability. ERP is mainly for manufacturing industry. However, the principles of ERP can also be applied to container terminal. We have clustered the workflow of container terminal, and analyzed the business process to generate the best workflow. The integrated database is designed to eliminate redundancy and keep integration. The core of ERP for container terminal is the planning facility such as berth planning and yard planning. The planning capability is very tightly coupled with data flow from client entities such as shipping companies. The ERP can handles the existing problems of container terminal operation that are mainly caused by lack of integration of whole of information resource in container terminal, ad-hoc and poor planning capability, disconnected and incorrect data from client companies. ERP approach not only van resolve the problems of container terminals but also promote adoption of information systems for container terminals in the world that have not yet implemented terminal operation systems.

Cyclodextrin Glucanotransferase와 Cyclodextrinase를 생산하는 Bacillus 속 세균의 분리와 그 효소들의 특성

권현주,남수완,김광현,곽영규,김병우 한국미생물생명공학회 ( 구 한국산업미생물학회 ) 1996 한국미생물·생명공학회지 Vol.24 No.3

토양으로부터 CGTase와 CDase를 함께 분비·생산하는 내열성 세균을 분리하였으며, 동정결과 Bacillus stearothermophilus로 판명되어 KJ16으로 명명하였다. Bacillus stearothermophilus KJ16 균주는 회분배양할 경우 2단계 성장을 하며 일차증식기에 CGTase를 생산하고 이차증식기에 CDase를 생산하는 양상을 보였다. 배치 pH를 7.0으로 조절하면서 배양할 경우, CGTase는 균의 증식에 따라 증가하고 CDase는 CGTase보다 뒤에 생산되었다. 그 생산양은 pH 비조절 배양에 비해 CGTase는 최대 12배(1300 mU/ml)까지, CDase는 2배(225 mU/ml)정도 증가되었다. 분리균주가 생산하는 CGTase는 분자량이 65,000 dalton으로 pH 6.0, 60℃에서 최적활성을 보이며 HPLC 분석을 통해 α-, β-, γ-CD를 42: 46: 12의 비율로 생산하는 α/β 혼합형 CGTase였다. 또한 CDase는 분자량이 68,000 dalton으로 pH 6.0, 55℃에서 최적활성을 보이며 최적기질은 γ-CD였다. A bacterium named as Bacillus stearothermophilus KJ16. The growth of the isolated strain occurred in two steps, and syntheses of CGTase and CDase were dependent on the growth cycle of the cell. CGTase was constitutively synthesized during the 1st growing phase, while CDase was synthesized inducibly during the 2nd growing phase. When the medium pH was controlled at 7.0, the maximun enzyme activities of CGTase and CDase were increased by 12-fold(1300 mU/ml) and 2-fold (225 mU/ml), respectively, compared with the pH-uncontrolled batch culture. The CGTase of the isolate converted soluble starch to CDs with the ratio of α-CD: β-CD: γ-CD=42:46:12 at 55℃. The optimal pH and temperature of CGTase were 6.0 and 60℃, respectively and the optimal pH and temperature of CDase were 6.0 and 55℃. The molecular weights of the purified CGTase and CDase were estimated to be 65,000 and 68,000 dalton, respectively. Among serveral substrates, γ-CD was most rapidly hydrolyzed by the purified CDase.

산업효소의 효모 표면발현(yeastsurfacedisplay)과 그 응용성

김현철,남수완 동의대학교 산업기술개발연구소 2006 産業技術硏究誌 Vol.20 No.-

최근 새로운 표면발현 시스템으로서 진핵세포인 효모 Saccharomyces cerevisiae를 이용하려는 시도가 활발히 진행되고 있다. 제빵효모 S. cerevisiae는 GRAS(generally recognized as safe)이며 오랫동안 많은 전통 생물공학산업에 이용되어 왔기 때문에 분자유전학 연구가 많이 되어 왔고 또 세포의 고농도 발효기술이 축적되어있다. 뿐만 아니라 효모는 진핵세포로서 고등생물과 유사한 유전자 구조 및 glycosylation을 포함하는 번역후 단백질 수식(post-translational modification) 과정을 가지기 때문에 고등세포 유래의 단백질을 활성상태로 발현할수 있다. 또한 기존의 bacteriophage의 경우 고밀도 표면발현이 가능한 경우는 약 50-60 amino acid로서 display할 수 있는 단백질의 크기에 한계가 있으며 대장균이나 그람양성세균의 경우에는 효소와 같은 분자량이 큰 단백질을 display할 수 있으나 표면발현의 밀도가 매우 낮은 문제점 등을 보완할수 있어서 많은 기능성 효소가 효모에서 표면발현 되었고 그 기능성을 확인하였다. 본 내용은 cycloinulooligosaccharide fructanotransferase와 endoinulinase를 효모 표면에 발현시켜 기능성 생물소재를 생산함을 확인하였다.