고열충격반응에 영향을 미치는 초파리 hsp70 유전자의 DNA 염기배열

이영훈,전은순,박충웅 ( Young Hoon Lee,Eun Soon Jeon,Chung Ung Park ) 생화학분자생물학회 1988 BMB Reports Vol.21 No.4

The DNA sequences required for heat shock regulated expression were examined in Drosophila melanogaster. Various hsp70-YP1 fusion constructs where D. melanogaster hsp70 gene and YP1 gene were used as a heat shock promoter and as a target gene, respectively, were transfected into D. melanogaster Schneider line 2 cells and their heat inducibility was determined by Northern analysis. The sequences containing two HSEs between -89 and -38 of the hsp70 were not sufficient to induce YP1 RNA upon heat shock whereas the sequences between -89 and +34 including the TATA box and the transcription start region of the hsp70 besides the HSEs were able to drive YP1 transcription during heat shock. The small deletion of the sequences near the transcription start region of the YP1 gene was also observed to repress the heat induced transcription of YPl RNA by the sequences between -89 and +34 of the hsp70.

고열충격반응에 영향을 미치는 초파리 hsp70 유전자의 DNA 염기배열

이영훈,전은순,박충웅,Lee, Young-Hoon,Jeon, Eun-Soon,Park, Chung-Ung 생화학분자생물학회 1988 한국생화학회지 Vol.21 No.4

The DNA sequences required for heat shock regulated expression were examined in Drosophila melanogaster. Various hsp70-YP1 fusion constructs where D. melanogaster hsp70 gene and YPI gene were used as a heat shock promoter and as a target gene, respectively, were transfected into D. melanogaster Schneider line 2 cells and their heat inducibility was determined by Northern analysis. The sequences containing two HSEs between -89 and -38 of the hsp70 were not sufficient to induce YP1 RNA upon heat shock whereas the sequences between -89 and +34 including the TATA box and the transcription start region of the hsp70 besides the HSEs were able to drive YP1 transcription during heat shock. The small deletion of the sequences near the transcription start region of the YP1 gene was also observed to repress the heat induced transcription of YP1 RNA by the sequences between -89 and +34 of the hsp70. 초파리 Drosophila melalnogaster에서 고열충격 유발을 조절하는데 필요한 DNA 염기배열을 조사하였다. hsp 70 유전자를 고열충격 촉진자로, YP1 유전자를 표적유전자로 하여 hsp70-YP1 융합유전자를 제조하고 이들 융합유전자를 이용하여 초파리 Schneider line 2 세포를 형질변환시키고 Northern 분석을 함으로써 hsp70-YP1 융합유전자의 고열충격 유발성을 결정하였다. 두개의 HSE (heat shock element) 가 존재하는 hsp70 유전자의 -89에서 -38까지 염기배열만으로는 고열충격에 의해 YP1 RNA를 유발시킬 수 없었지만 두 개의 HSE 이외에 hsp70 유전자의 TATA 박스와 전사시작점을 포항하고 있는 -89에서 +34까지 염기배열은 고열충격에 의해 YP1 RNA를 유발시킬 수 있었다. 또한 YP1 유전자의 전사시 작정 근처의 염기배열이 조금 결실되었을 때 hsp70 유전자의 -89에서 +34까지 염기배열에 의한 YP1 RNA의 고열충격 유발이 억제되는 것이 관찰되었다.

Escherichia coli M1 RNA 유전자의 전사 종결

이영미,이영훈,박충웅,Lee, Young-Mi,Lee, Young-Hoon,Park, Chung-Ung 생화학분자생물학회 1989 한국생화학회지 Vol.22 No.3

Escherichia coli RNase P의 RNA 성분인 M1 RNA를 코드하는 유전자에 연속적으로 존재하는 종결제 T1, T2, T3의 효율을 측정하였다. 이를 위하여 M1 RNA 유전자의 3'-부분에 있는 염기배열을 차례로 결실시키고, 결실된 염기배열을 갖는 M1 RNA 유전자를 Gal K 발현 벡타인 pKO100에 클로닝하여 갈락토키나제 활성도를 측정하였다. 주된 종결반응은 첫번째 종결제인 T1에서 94%의 효율로 일어났으며, 세 개의 종결제가 모두 존재할 때는 거의 100% 효율로 종결되었다. 또한 이러한 종결제를 통과하여 전사된 RNA가 가지는 ORF (open reading frame)들에서 만들어 질 수 있는 폴리펩티드에 대해서도 검토하였다. The deletion experiment at the 3´flanking sequences of the gene for M1 RNA, the RNA component of Echerichia coli ribonuclease P, was carried out to estimate the efficiency of tandem terminators, T1, T2, and T3. The serially deleted sequences of the M1 RNA gene were cloned into GalK expression vector pK0100 and relative galactokinase activities were measured. The major termination occurred at the first terminator, T1, and three terminators all together functioned with almost 100% efficiency. Biosynthesis of the polypeptides encoded by readthrough transcripts from the terminators was also discussed.

Escherichia coli M1 RNA 유전자의 전사 종결

이영미,이영훈,박충웅 ( Young Mi Lee,Young Hoon Lee,Chung Ung Park ) 생화학분자생물학회 1989 BMB Reports Vol.22 No.3

The deletion experiment at the 3` flanking sequences of the gene for M1 RNA, the RNA component of Escherichia coli ribonuclease P, was carried out to estimate the efficiency of tandem terminators, T1, T2, and T3. The serially deleted sequences of the M1 RNA gene were cloned into GalK expression vector pK0100 and relative galactokinase activities were measured. The major termination occurred at the first terminator, T1, and three terminators all together functioned with almost 100% efficiency. Biosynthesis of the polypeptides encoded by readthrough transcripts from the terminators was also discussed.

대장균 rnpA 돌연변이를 보완작용할 수 있는 Brevibacterium albidum 염색체 부위의 특성

이영미,채건상,박충웅,이영훈 ( Young Mi Lee,Keon Sang Chae,Chung Ung Park,Young Hoon Lee ) 생화학분자생물학회 1990 BMB Reports Vol.23 No.4

The chromosomal region containing the gene capable of complementing the E. coli rnpA mutation was obtained from genomic DNA of Brevibacterium albidum. The selection procedure was based on the formation of an enzymatically functional hybrid RNase P enzyme by heterologous gene products in E. coli. Unlike the E. coli rnpB gene, the B. albidum DNA region on a multicopy plasmid did not complement the E. coli rnpB mutation. The deletion analysis indicates that the rnpA-complementing activity resides in the 2.75 kb region.

새로운 LacZ Gene Fusion 및 Gene Cartridge Vector의 개발에 관한 연구

박교선,이영훈,박충웅,최창진 圓光大學校 基礎自然科學硏究所 1987 基礎科學硏究誌 Vol.6 No.2

In this study, a new lacZ gene vector, plasmid PGEM3-lacZ was constucted, which could be used for the study of the gene regulation and the function of proteins. Plasmid pMC1871 was digested with restriction endonuclease Pst I and Sma I to obtain a DNA fragment with the lacZ-gene(3091 bp). Recombinant plasmid pGEM3-lacZ was constructed by inserting the lacZ gene DNA(Pst I-Sma I cut DNA; 3091 bp) of pMC1871 into the Pst I and Hinc Ⅱ sites of plasmid pGEM3. After then, Recombinant plasmid pGEM3-lacZ DNA was transformated into E. coli HB 101. pGEM3-lacZ DNA/HB 101 was screened on the NZY plate with ampicillin and X-gal(5-Bromo-4-Chloro-3-Indolyl-β-D-Galactoside) by selecting blue colonies. The restriction map of pGEM.3-1acZ DNA was constructed by the following restriction enzymes :Bgl I, Xbal I, Bam HI, EcoR I, Hind Ⅲ, Kpn I, Pst I, Sma Ⅰ, Sal Ⅰ. From the restriction map, the lacZ DNA was confirmed to be present at the expected position in the plasmid pGEM3-lacZ. pGEM3-lacZ/HB101 gives about 10 times more lacZ DNA than pMC 1871/HB101.

rnpB 유전자 발현에 RNA 이차구조가 미치는 영향

전은순,이영훈,박충웅,조인호 全北大學校 基礎科學硏究所 1988 基礎科學 Vol.11 No.1

E. coli rnpB gene의 발현에 영향을 미치는 M1 RNA structural gene의 부분을 조사하였다. P-1 promoter 에서 합성되는 RNA의 5' 부분이 이루는 이차구조를 분석한 결과 +1 에서 부터 +72 까지에 존재하는 hairpin loop 구조가 rnpB gene의 발현에 중요한 역할을 하는 것으로 밝혀졌다.

성장속도에 따른 대장균 rnpB 유전자 발현의 조절작용

이영훈,정재훈,박충웅,이석재,전은순,이학란,정영환,조명선 한국유전학회 1993 Genes & Genomics Vol.15 No.1

Growth rate-dependent expression of the rnpB gene encoding M1 RNA, the RNA component of Escherichia coli RNase P, was examined using a hybrid transducing λ phage. The hybrid transducing λ phage was constructed carrying the lacZ gene fused to the promoter for the rnpB gene. The amounts of β-galactosidase were analyzed in the E. coli lysogen carrying this transducing phage grown in different growth media to see dependence of the rnpB transcription on growth rate. The synthesis of β-galactosidase from the rnpB-lacZ fusion increased with growth rate. This result suggests that the rnpB promoter activity is dependent on the growth rate.

대장균에서 많은 수의 플라스미드에 존재하는 rnpB 유전자 전사의 성장기의 따른 조절 작용

이영훈,박충웅,전은순,정영환,길민찬,은영아 한국유전학회 1993 Genes & Genomics Vol.15 No.2

The Escherichia coli rnpB gene encodes M1 RNA, the catalytic component of RNase P which is a processing enzyme for tRNA maturation. Biosynthesis of M1 RNA is known to be stringently regulated. The rnpB transcription under different growth phases of E. coli was examined using the truncated rnpB in the multicopy plasmids which could generate the metabolically unstable M1 RNA transcript. Biosynthesis of this RNA was directly analyzed during the transition from the exponential phase of growth to the stationary phase. During the transition the rnpB transcription became repressed. The discriminator motif CGCC adjacent to the transcription start-point of the rnpB gene way altered to the TATT using site-directed mutagenesis. The modified promoter containing the mutated discriminator was at least partially rescued from this growth phase-dependent repression of the rnpB transcription. The results clearly show that the discriminator motif play a key role in control of the rnpB transcription under different growth phases.

성장속도에 따른 대장균 rnpB 유전자 발현의 조절작용

전은순,이학란,이석재,정영환,조명선,정재훈,이영훈,박충웅 全北大學校 基礎科學硏究所 1994 基礎科學 Vol.16 No.-

Growth rate-dependent expression of the rnpB gene encoding M1 RNA, the RNA component of Escherichia coli RNase P, was examined using a hybrid transdcuing λ phage. The hybrid transducing λ phage was constructed carrying the lacZ gene fused to the promoter for the rnpB gene. The amounts of β-galactosidase were analyzed in the E. coli lysogen carrying this transducing phage grown in different growth media to see dependence of the rnpB transcription on growth rate. The synthesis of β-galactosidase from the rnpB-lacZ fusion increased with growth rate. This result suggests that the rnpB promoter activity is dependent on the growth rate.