대장균 변이주 내에서 λ P L Promoter 를 갖는 재조합 트립토판 유전자의 발현과 안정성

허태린,김현수,이세영 ( Tae Lin Huh,Hyeon Soo Kim,Se Yong Lee ) 생화학분자생물학회 1987 BMB Reports Vol.20 No.1

We studied the effects of trpR and tnaA gene in E. coli mutants on the expression level and stability of recombinant trp plasmids, pPLT and pPLPT. In the absence of c1857 gene, recombinant plasmid pPLPT, mainly express its trp gene by trp promoter with the aids of λ P_L promoter, showed 1.5 times higher activities of tryptophan synthetase than those of trp gene in plasmid pPLT which express its trp gene by λ P_L promoter only. In contradiction to its elevated trp gene expression, plasmid stability was more decreased than that of plasmid pPLT. In tnaA^+ host, trp gene expressions and stabilities of recombinant trp plasmids, pPLPT in tnaA^- host. In the presence of plasmid pYS46 which conferrs cI857 gene, expression levels of trp plasmids, pPLT were higher than those of plasmid pPLPT. At that time, plasmid pY S46 showed very poor plasmid stability in its host.

고등동물의 Pyruvate dehydrogenase complex

허태린 한국분자생물학회 1992 분자생물학뉴스 Vol.4 No.1

일반특강 및 구두발표 논문 초록 ; E . coli 숙주변이주내 재조합 trp+ 플라스미드의 안정성증대

허태린,강충민,이세영 한국생화학분자생물학회 (구 한국생화학회) 1987 생화학분자생물학회 추계학술발표논문집 Vol.- No.-

대장균 변이주 내에서 ${\lambda}P_L$ Promoter를 갖는 재조합 트립토판 유전자의 발현과 안정성

허태린,김현수,이세영,Huh, Tae-Lin,Kim, Ryeon-Soo,Lee, Se-Yong 생화학분자생물학회 1987 한국생화학회지 Vol.20 No.1

재조합 플라스미들은 pPLT와 pPLPT 내에 있는 트립토판 유전자(trp)들의 발현과 안정성에 미치는 숙주세포들의 trpR 그리고 tnaA 유전자의 영향에 대하여 조사하였다. 숙주세포가 cI857 유전자를 갖고 있지 않을 경우, ${\lambda}P_L$과 함께 trp promoter에 의해 발현되는 재조합 trp 유전자를 갖고 있는 플라스미드 pPLPT는 ${\lambda}P_L$ promoter만에 의해서 발현되는 재조합 trp 유전자를 갖고있는 플라스미드 pPLT 보다 트립토판 유전자의 발현정도가 1.5배 이상 높게 나타났으나 플라스미드의 안정성은 오히려 저하되었다. 또한 숙주세포가 $tnaA^+$ 일 때보다 $tnaA^-$일 때 재조합 trp 유전자의 발현정도는 더 낮았으며 안정성도 감소되었다. 그러나 숙주세포 내에 cI857 유전자가 재조합 된 플라스미드 pYS46이 존재시 trp 유전자의 발현도는 플라스미드 pPLT에서 더 높게 나타났으며 이 때 플라스미드 pYS46 의 안정성은 매우 낮게 나타났다. We studied the effects of trpR and tnaA gene in E. coli mutants on the expression level and stability of recombinant trp plasmids, pPLT and pPLPT. In the absence of cI857 gene, recombinant plasmid pPLPT, mainly express its trp gene by trp promoter with the aids of ${\lambda}P_L$ promoter, showed 1.5 times higher activities of tryptophan synthetase than those of trp gene in plasmid pPLT which express its trp gene by ${\lambda}P_L$ promoter only In contradiction to its elevated trp gene expression, plasmid stability was more decreased than that of plasmid pPLT. In $tnaA^+$ host, trp gene expressions and stabilities of recombinant trp plasmids, pPLPT in $tnaA^-$ host. In the presence of plasmid pYS46 which conferrs cI857 gene, expression levels of trp plasm ids, pPLT were higher than those of plasmid pPLPT. At that time, plasmid pYS46 showed very poor plasmid stability in its host.

Zebrafish Model을 이용한 척추동물의 발생연구

허태린 한국과학기술원 유전공학연구소 1996 생명공학동향 Vol.4 No.4

Trichoderma viride 섬유소 분해효소에 관한 연구 ( 1 ) Carboxymethylcellulose 에 의해서 섬유소 분해효소들의 성질

허태린,이세영 ( Tae Lin Huh,Se Yong Lee ) 생화학분자생물학회 1981 BMB Reports Vol.14 No.1

In order to investigate the characteristics of cellulases induced by carboxymethylcellulose, Trichoderma viride QM 9414 was cultured on CMC. The culture filtrates were concentrated by (NH₄)₂SO₄ fractionation and used as enzyme sources, The enzyme preparations were further purified by gel filteration and DEAF-Sephadex chromatography and assayed for cotton activity, CMC activity and β-glucosidase activity. The following results were obtained: 1. The optimum pH of the cotton activity, CMC activity and β-glucosidase activity were between pH 4. 8-5. 2. 2. The optimum temperature of the cotton activity was 50℃ and of CMC activity 55℃, but the optimum temperature of the β-glucosidase activity was 40℃. 3. Cellulose induced by CMC contained high β-glucosidase activity compared with cellulase induced by Avicel and cellobiose. 4. A low-molecular weight CMCase activity was separated from the rest by Sephadex G-75 gel and most of β-glucosidase activity was spearated by Bio P-150 column chromatography. The remaining β-glucosidase was separated from CMC ass by DEAE-Sephadex chromatography. From these, a cellulase that could only degrade CMC and cellobiose but not crystalline cellulase were obtained. 5. Trichoderma cellulases induced by CMC could hydrolyze cotton, CMC, and cellobiose into glucose. 6. CMCase has both endoglucanase and exoglucanase activity which degrade cellooligesaccharides and cellobiose readily into glucose in the absence of β-glucosidase, 7. On the basises of results obtained from this study a mechanism of Trichoderma viride cellulases is proposed.

대장균 내에서 λ P L Promoter 를 갖는 재조합 트립토판 유전자들의 구성과 발현

허태린,김현수,이세영 ( Tae Lin Huh,Hyeon Soo Kim,Se Yong Lee ) 생화학분자생물학회 1987 BMB Reports Vol.20 No.1

For the elevation of the level of tryptophan biosynthetic gene (trp) expression, we cloned trpL(△ att) trpEDCBA gene which was resistant to partial termination of transcription in the presence of excess tryptophan in the culture medium. When trp1,(0 att) trpEDCBA gene on λ P_L promoter containing plasmid p a 8 was introduced into E. coli which confers a temperature sensitive c1857 gene on its chromosome, activity of tryptophan synthetase expressed by λ P_L promoter was 3 times higher than that of tryptophan synthetase which was expressed by trp promoter.

고등동물 Isocitrate Dehydrogenase Isoenzyme 들의 유전자 구조와 기능

허태린 경북대학교 생명공학연구소 1996 遺傳工學硏究所報 Vol.11 No.1

N/A

대장균 내에서 ${\lambda}P_L$를 Promoter 갖는 재조합 트립토판 유전자들의 구성과 발현

허태린,김현수,이세영,Huh, Tae-Lin,Kim, Hyeon-Soo,Lee, Se-Yong 생화학분자생물학회 1987 한국생화학회지 Vol.20 No.1

대장균 내에서 트립토판 생합성 유전자(trp)의 발현증대를 위하여 attenuator가 결실되어 있어서 배지중에 과량의 트립토판이 존재시에도 trp 유전자의 transcnption이 partial termination되지 않도록 변형된 trpL(${\Delta}$ att) trpEDCBA 유전자를${\lambda}P_L$ promoter를 갖고 있는 플라스미드 $p{\lambda}8$에 재조합하여 대장균 내에서 발현시켰다. 구성된 재조합 trp 플라스마드들을 온도감수성 cI857 유전자를 갖고 있는 대장균 숙주세포 내에서 발현시켰을 때 ${\lambda}P_L$ promoter에 의해서 발현된 tryptophan synthetase의 역가가 trp promoter에 의해 발현될 때보다 3 배 이상 증가되었다. For the elevation of the level of tryptophan biosynthetic gene (trp) expression. we cloned trpL(${\Delta}$att)trpEDCBA gene which was resistant to partial termination of transcription in the presence of excess tryptophan in the culture medium. When trpL(${\Delta}$ att) trpEDCBA gene on ${\lambda}P_L$ promoter containing plasmid $p{\lambda}8$ was introduced into E. coli which confers a temperature sensitive cI857 gene on its chromosome. activity of tryptophan synthetase expressed by ${\lambda}P_L$ promoter was 3 times higher than that of tryptophan synthetase which was expressed by trp promoter.

Cellulases of Trichoderma viride (I) - Induction of cellulases by carboxymethylcellulose and their mode of action

허태린,이세영,Huh, Tae-Lin,Lee, Se-Yong 생화학분자생물학회 1981 한국생화학회지 Vol.14 No.1

섬유소 분해효소 생성균주인 Trichoderma viride QM9414를 CMC에서 배양시 생성된 섬유소 분해효소들의 성질과 cotton, carboxymethylcellulose(CMC) 그리고 cellobiose 분해 기작을 조사하였으며, culture filtrate를 $(NH_4)_2SO_4$ fractionation, gel filtration과 DEAE-Sephadex column chromatography를 통해 cotton 분해력, CMC 분해력 그리고 cellobiose 분해력의 분리를 시도하였다. 얻어진 결과는 다음과 같았다. 1. Cotton activity, CMC activity, 그리고 ${\beta}$-glucosidase의 최적 활성 pH는 4.8-5.2로 나타났다. 2. Cotton activity의 최적활성온도는$50^{\circ}C$이며 CMC activity는 $55^{\circ}C$로 나타났고 ${\beta}$-glucosidase는 $40^{\circ}C$ 최적활성 온도로 나타났다. 3. CMC에 의해서 유도된 섬유소 분해효소는 결정형 섬유소나 cellobiose에 의해서 유도된 섬유소 분해효소와는 달리 높은 ${\beta}$-glucosidase를 보였다. 4. CMC에 의해서 유도된 섬유소 분해 효소 농축액을 Sephadex G-75 gel에 통과시켰을 때 고분자 섬유소 분해효소와 CMC에 대해서만 낮은 역가를 보여주는 저분자 CMCase가 분리되었으며 이 고분자 섬유소 분해 효소 분획을 다시 Bio Gel P-150에 통과시킨 결과 대부분의 ${\beta}$-glucosidase와 cotton 분해 효소가 CMC 분해 효소로 부터 분리되었다. CMC 분해 효소 분획은 DEAE Sephadex A-50 이온교환수지에 재통과시켜 잔여 ${\beta}$-glucosidase를 제거하고 CMC 분해효소만 단리하였다. 5. T. viride QM9414로부터 CMC에 의해서 유도된 섬유소 분해효소 분획들은 모두 cotton과 CMC를 분해하여 glucose와 cellobiose를 생성하였는데 최종 산물은 주로 glucose 였다. 6. CMCase는 endoglucanase와 exoglucanase 활성을 둘다 갖고 있는 것처럼 보였고 CMC와 cellobiose로부터 ${\beta}$-glucosidase 존재 여부와 관계없이 glucose를 생성하였다. 7. 이 연구의 결과를 종합하여 Trichoderma viride cellulase의 cellulose 분해 작용 기작을 제안하였다. In order to investigate the characteristics of cellulases induced by carboxymethylcellulose, Trichoderma viride QM 9414 was cultured on CMC. The culture filtrates were concentrated by $(NH_4)_2SO_4$ fractionation and used as enzyme sources. The enzyme preparations were further purified by gel filteration and DEAF-Sephadex chromatography and assayed for cotton activity, CMC activity and ${\beta}$-glucosidase activity. The following results were obtained: 1. The optimum pH of the cotton activity, CMC activity and ${\beta}$-glucosidase activity were between pH 4.8-5.2. 2. The optimum temperature of the cotton activity was $50^{\circ}C$ and of CMC activity $55^{\circ}C$, but the optimum temperature of the ${\beta}$-glucosidase activity was $40^{\circ}C$. 3. Cellulose induced by CMC contained high ${\beta}$-glucosidase activity compared with cellulase induced by Avicel and cellobiose. 4. A low-molecular weight CMCase activity was separated from the rest by Sephadex G-75 gel and most of ${\bata}$-glucosidase activity was spearated by Bio P-150 column chromatography. The remaining ${\beta}$-glucosidase was separated from CMC ase by DEAE-Sephadex chromatography. From these, a cellulase that could only degrade CMC and cellobiose but not crystalline cellulase were obtained. 5. Trichoderma cellulases induced by CMC could hydrolyze cotton, CMC, and cellobiose into glucose. 6. CMCase has both endoglucanase and exoglucanase activity which degrade cellooligesaccharides and cellobiose readily into glucose in the absence of ${\beta}$-glucosidase. 7. On the basises of results obtained from this study a mechanism of Trichoderma viride cellulases is proposed.