염주말(Chaetomorpha moniligera)의 5.8S rRNA의 일차구조 및 이차구조 결정

정태능,박인원,Johng, Tae-Neung,Park, In-Won 생화학분자생물학회 1988 한국생화학회지 Vol.21 No.1

The primary structure of 5.8S rRNA from Chaetomorpha moniligera, one of green algae, was determined by enzymatic method. It consists of 158 nucleotides and contains a 2'-O-methylated purine nucleoside at the position of 42 and a 2'-O-methylated guanosine at the position of 76. The 5.8S rRNA from Chaetomorpha moniligera has 80% and 78% sequence homology with those of Chlamydomonas reinhardii, another green algae, and of yeast, respectively. We have constructed the possible secondary structure of S.8S rRNA from Chaetomorpha moniligera and we have found that the constructed secondary structure fits very well for the formation of complex with 26S rRNA from yeast as was proposed by Ursi et al (1983). These findings suggest that the sequences of eukaryotic 5.8S rRNA are well conserved during the evolution. 녹조류의 일종인 염주말의 5.8S rRNA의 일차구조를 효소적 방법으로 결정하였다. 염주말의 5.8S rRNA는 158개의 누클레오티드로 구성되어 있으며, 42번 위치에는 2'-O-메틸화된 푸린 누클레오시드를, 76번 위치에는 2'-O-메틸화된 구아노신을 함유하고 있다. 염주말의 5.8S rRNA는 다른 한 단세포 녹조류인 Chlamydomonas reinhardii의 것과는 80%의 상동성을 가지고 있고 효모의 5.8S rRNA와는 78%의 상동성을 가지고 있다. 다른 진핵세포의 5.8S rRNA의 일차구조들과 비교하여 보존된 연속부분들에 기초하여 염주말의 5.8S rRNA의 이차구조를 구성해보았다. 구성한 이차구조는 Ursi 등 (1983)이 제안한 것과 같은 26S rRNA-5.8S rRNA 복합체를 형성할 수 있는 그러한 것이었다.

염주말 ( Chaetomorpha moniligera ) 의 5 . 8 S rRNA 의 일차구조 및 이차구조 결정

정태능,박인원 ( Tae Neung Johng,In Won Park ) 생화학분자생물학회 1988 BMB Reports Vol.21 No.1

The primary structure of 5.8S rRNA from Chaetomorpha moniligera, one of green algae, was determined by enzymatic method. It consists of 158 nucleotides and contains a 2`-O-methylated purine nucleoside at the position of 42 and a 2`-O-methylated guanosine at the position of 76. The 5.8S rRNA from Chaetomorpha moniligera has 80% and 78% sequence homology with those of Chlamydomonas reinhardii, another green algae, and of yeast, respectively. We have constructed the possible secondary structure of 5.8S rRNA from Chaetomorpha moniligera and we have found that the constructed secondary structure fits very well for the formation of complex with 26S rRNA from yeast as was proposed by Ursi et al (1983). These findings suggest that the sequences of eukaryotic 5.8S rRNA are well conserved during the evolution.

녹조류 Chaetomorpha moniligera 의 5S rRNA의 누클레오티드 결합 순서와 이차구조

정태능,정연희,박인원 ( Tae Neung Johng,Yean Hee Joung,In Won Park ) 생화학분자생물학회 1987 BMB Reports Vol.20 No.1

The nucleotide sequence of 5S rRNA from a multicellular green alga Chaetomorpha moniligera was determined. The 5S rRNA is composed of 120 nucleotides and does not have any modified base. The nucleotide sequence of the 5S rRNA has wide range of similarities to the other green algae (53-71%). In spite of this, the secondary structure of the 5S rRNA has the general eukaryotic characteristics. Most of its bases in single-stranded regions are well conserved like in other eukaryotic 5S rRNA. v

염주말에 있어서의 rRNA 유전자들의 배열

정태능,김동수,임자혜,박인원 ( Tae Neung Johng,Dong Soo Kim,Ja Hei Ihm,In Won Park ) 생화학분자생물학회 1992 BMB Reports Vol.25 No.1

Using fragments of appropriate sizes of 5.8S, 17S, and 25S rRNAs as probes, the arrangement of rRNA genes of cytoplasmic rDNA of Chaetomorpha moniligera has been determined, and also the restriction map of the rDNA has been analyzed. The length of the repeating unit of rDNA is approximately 17 kb long and its organization is as follows: (17S rDNA)-ITS 1-(5.8S rDNA)-ITS 2-(25S rDNA). It has been found that 5S rDNA is not present within 17 kb repeating unit. Interestingly, the presence of an intron of about 0.5 kb has been found within the region of 25S rDNA.

The Nucleotide Sequence and the Secondary Structure of 5S rRNA from a Green Alga Chaetomorpha moniligera

정태능,정연희,박인원,Johng, Tae-Neung,Joung, Yean-Hee,Park, In-Won Korean Society for Biochemistry and Molecular Biol 1987 한국생화학회지 Vol.20 No.1

The nucleotide sequence of 5S rRNA from a multicellular green alga Chaetomorpha moniligera was determined. The 5S rRNA is composed of 120 nucleotides and does not have any modified base. The nucleotide sequence of the 5S rRNA has wide range of similarities to the other green algae (53-71%). In spite of this, the secondary structure of the 5S rRNA has the general eukaryotic characteristics. Most of its bases in single-stranded regions are well conserved like in other eukaryotic 5S rRNA. 녹조류의 하나인 Chaetomorpha moniligera의 5S rRNA의 누클레오티드의 결합 순서를 결정하였다. 이 5S rRNA는 120개의 누클레오티드로 구성되어 있으며, 변형된 염기가 하나도 없었다. 이 5S rRNA의 누클레오티드의 결합 순서는 그밖의 녹조류의 5S rRNA들파 53%에서 71%까지의 광범위한 유사성을 가지고 있다. 그럼에도 불구하고 이 5S rRNA의 이차구조는 일반적인 진핵세포의 특성을 가지고 있다. 단일 가닥으로 된 부분에 있는 염기들은 그밖의 진핵세포의 것들과 마찬가지로 잘 보존되어 있다.

불순한 국산 $^{32}P-H_3PO_4$의 정제와 [${\gamma}-^{32}P$]-ATP의 합성에의 응용

박인원,정태능,Park, In-Won,Johng, Tae-Neung 생화학분자생물학회 1984 한국생화학회지 Vol.17 No.1

이 연구의 목적은 한국 에너지 연구소에서 생산하는 불순한 $^{32}P-H_3PO_4$를 생화학적 용도에 알맞는 순도까지 정제하고, 그것을 [${\gamma}-^{32}P$]-ATP의 합성에 응용하고자 하는 데 있다. $^{32}P-H_3PO_4$의 조용액에 들어 있는 주되는 불순물들은 황산 이온과 나트륨 이온 들임을 알았다. 양이온 교환수지와 음이온 교환수지들의 두 층으로 된 배합 대롱을 사용하여 이 이온들을 불순한 $^{32}P-H_3PO_4$ 용액에서 제거하였다. 이러한 방법으로 불순한 $^{32}P-H_3PO_4$를 생화학적 순도까지 정제하였다. $^{32}P-H_3PO_4$의 회수율은 약 70%이었다. 정제된 $^{32}P-H_3PO_4$는 [${\gamma}-^{32}P$]-ATP의 합성에 이용가능할 정도로 충분히 순수하였다. 합성한 [${\gamma}-^{32}P$]-ATP는 RNA 조각들의 5' 말단의 히드록시기를 표지하는 데 효과적으로 이용되었다. The purpose of this research was, primarily, to develop the methods for the purification of the crude $^{32}P-H_3PO_4$ produced by the Korean Energy Research Institute to biochemical purity, and then, to apply the purified $^{32}P-H_3PO_4$ for the synthesis of [${\gamma}-^{32}P$]-ATP. The major impurities contained in the crude solution of $^{32}P-H_3PO_4$ were sulfate and sodium ions. These ions were eliminated from the crude $^{32}P-H_3PO_4$ solution by employing the combined column consisting of two layers of cation-and anion-exchangers. In this way, the crude $^{32}P-H_3PO_4$ was purified to biochemical purity. The recovery of $^{32}P-H_3PO_4$ was about 70%. The purified $^{32}P-H_3PO_4$ thus obtained was sufficiently pure for the synthesis of [${\gamma}-^{32}P$]-ATP, which was effectively used for the labeling of the 5'-OH termini of RNA fragments.

[${\gamma}-^{32}P$]-ATP 합성의 수득률 증가와 그 정제를 위한 방법 연구

고문주,정태능,박인원,Koh, Moon-Joo,Johng, Tae-Neung,Park, In-Won 생화학분자생물학회 1986 한국생화학회지 Vol.19 No.4

국산 $[^{32}P]-H_3PO_4$ 조용액으로부터 [${\gamma}-^{32}P$]-ATP를 합성할 때 그 수득률을 증가시킬 수 있는 방법과 합성된 [${\gamma}-^{32}P$]-ATP의 정제법을 개량하기 위한 방법을 고안해 보았다. 합성 단계에서는 반응 물질과 효소의 농도를 일정하게 유지하면서 반응 용액의 부피를 증가시키는 방법이 [${\gamma}-^{32}P$]-ATP의 합성을 증가시켰으며 정제 단계에서는 DEAE-Sephadex A-25 이온 교환 수지를 이용하여 0.1 M부터 0.4 M까지의 TEAB 용액을 직선기울기로 농도를 변화시키면서 용리시키는 방법이 가장 효과적임을 알았다. We have designed a procedure for the improvement of yield of [${\gamma}-^{32}P$]-ATP synthesis starting from crude $[^{32}P]-H_3PO_4$. Increase of volume of reaction mixture at a constant ratio between the amount of enzymes and reactants increased the yield of [${\gamma}-^{32}P$]-ATP. The product was purified in a simple procedure using DEAE-sephadex A-25 ion exchanger and eluting the product with linear gradient concentration (0.1 M to 0.4 M) of triethylamine bicarbonate.

불순한 국산 32P - H3PO4 의 정제와 ( γ - 32P ) - ATP 의 합성에의 응용

박인원,정태능 ( In Won Park,Tae Neung Johng ) 생화학분자생물학회 1984 BMB Reports Vol.17 No.1

The purpose of this research was, primarily, to develop the methods for the purification of the crude ^(32)P-H₃PO₄ produced by the Korean Energy Research Institute to biochemical purity, and then, to apply the purified ^(32)P-H₃PO₄ for the synthesis of [γ-^(32)P]-ATP. The major impurities contained in the crude solution of ^(32)P-H₃PO₄ were sulfate and sodium ions. These ions were eliminated from the crude ^(32)P-H₃PO₄ solution by employing the combined column consisting of two layers of cation-and anion-exchangers. In this way, the crude ^(32)P-H₃PO₄ was purified to biochemical purity. The recovery of ^(32)P-H₃PO₄ was about 70%. The purified ^(32)P-H₃PO₄ thus obtained was sufficiently pure for the synthesis of [γ-^(32)P]-ATP, which was effectively used for the labeling of the 5`-OH termini of RNA fragments.

염주말의 핵에 있는 큰 소단위체 선구 rRNA 인트론의 구조

김동수,정연희,위호선,이승주,정태능,박인원 ( Dong Soo Kim,Yeon Hee Joung,Ho Sun Wee,Seung Joo Lee,Tae Neung Johng,In Won Park ) 생화학분자생물학회 1993 BMB Reports Vol.26 No.3

Recently, we have reported that the nuclear large subunit pre-rRNA of the green alga Chaetomorpha moniligera has an intron. Now we report that the intron in this pre-rRNA self-splices in vitro and that its structure constructed shows that it is one of the group I B introns. The self-splicing reaction of this intron is inhibited by gentamycin or L-arginine. The intron consists of 653 nucleotides. The primary structure of the intron was derived from the sequence of a region of 25S rDNA that contains the intron. Its 5` terminal base is cytosine and 3` terminal base of the 5` exon that is linked to the intron is uracil. On the other hand, on the basis of tripartite structure that is proposed to form for the selection of 3` splice site, 3` terminal base of the intron has been inferred to be guanine on the position 653. The 5` terminal base of the 3` exon that is linked to this guanine is uracil. Like the other group I introns, the intron in pre-rRNA of Chaetomorpha moniligera contains nine base-paired stems P1 through P9, and it contains also four conserved sequence elements in the following order: 5`-P(GAAUGCGGGAGC)-Q(CCUCCGCAGG)-R(CCCCUAGAGA-CUAGA)-S(AUUCGAGAGUCCU)-3`. Major distinct features of this intron differing from most of the other group I introns are that it has an extra stem P6c which is formed between 3` half of the stem P6a and 3` half of the stem P6, and that the stem P8 has a long extension which contains an open reading frame.

염주말의 핵에 있는 큰 소단위체 선구 rRNA 인트론의 구조

김동수,정연희,위호선,이승주,정태능,박인원,Kim, Dong-Soo,Joung, Yeon-Hee,Wee, Ho-Sun,Lee, Seung-Joo,Johng, Tae-Neung,Park, In-Won 생화학분자생물학회 1993 한국생화학회지 Vol.26 No.3

우리는 최근에 녹조류인 염주말의 핵에 있는 선구 rRNA가 인트론을 가지고 있다는 것을 보고한 바 있다. 우리는 여기서 이 선구 rRNA의 인트론이 시험관에서 자기이어맞추기를 하며, 이 인트론은 그 구조로 보아 I군 인트론의 B형에 속한다는 것을 보고하고자 한다. 이 인트론의 자기이어맞추기 반응은 겐타마이신 또는 L-아르기닌으로 억제된다. 이 언트론은 653개의 누클레오티드로 구성되어 있다. 우리는 인트론의 일차구조를 인트론을 함유하는 25S rDNA 부분의 염기 결합순서로부터 추정하였다. 인트론의 5' 말단의 염기는 시토신이고 그것과 연결된 5' 엑손의 3' 말단의 엽기는 우라실이다. 한편, 인트론의 3' 말단이 자기이어맞추기 자리를 선택할 때 형성하는 삼중복합체 구조로부터 추정한 인트론의 3' 말단의 엽기는 653번 위치에 있는 구아닌이고 그것과 연결된 3' 엑손의 5' 말단 염기는 우라실이다. 다른 I군 인트론들과 마찬가지로, 염주말의 선구 rRNA의 인트론은 Pl에서 P9까지 9개의 줄기를 가지며 4개의 보존된 연속부분 요소들을 다음과 갇은 순서로 함유하고 있다 :5'-P(GAAUGCGGGAGC)-Q(CCUCCGCAGG)-R(CCCCUAGAGACUAGA)-S(AUUCGAGAGUCCU)-3'. 이 인트론이 대부분의 다른 I군 인트론들과 주요하게 다른 점은 줄기 P6a의 3' 반쪽과 P6의 3' 반쪽 사이에 18개의 누클레오티드로 구성되는 가외의 줄기 P6c가 있다는 것과 줄기 P8이 open reading frame을 가지고 있고 크게 확대되어 있다는 것이다. Recently, we have reported that the nuclear large subunit pre-rRNA of the green alga Chaetomorpha moniligera has an intron. Now we report that the intron in this pre-rRNA self-splices in vitro and that its structure constructed shows that it is one of the group I B introns. The self-splicing reaction of this intron is inhibited by gentamycin or L-arginine. The intron consists of 653 nucleotides. The primary structure of the intron was derived from the sequence of a region of 25S rDNA that contains the intron. Its 5' terminal base is cytosine and 3' terminal base of the 5' exon that is linked to the intron is uracil. On the other hand, on the basis of tripartite structure that is proposed to form for the selection of 3' splice site, 3' terminal base of the intron has been inferred to be guanine on the position 653. The 5' terminal base of the 3' exon that is linked to this guanine is uracil. Like the other group I introns, the intron in pre-rRNA of Chaetomorpha moniligera contains nine base-paired stems P1 through P9, and it contains also four conserved sequence elements in the following order: 5'-P(GAAUGCGGGAGC)-Q(CCUCCGCAGG)-R(CCCCUAGAGA-CUAGA)-S(AUUCGAGAGUCCU)-3'. Major distinct features of this intron differing from most of the other group I introns are that it has an extra stem P6c which is formed between 3' half of the stem P6a and 3' half of the stem P6, and that the stem P8 has a long extension which contains an open reading frame.