sfs1 유전자의 cAMP - cAMP receptor protein 에 의한 발현 조절

최용락(Yong Lark Choi),유주순(Ju Soon Yoo),이승진(Seung Jin Lee),이희영(Hee Young Lee),정수열(Soo Yeol Chung) 한국응용생명화학회 1996 Applied Biological Chemistry (Appl Biol Chem) Vol.39 No.3

We have cloned several E. coli sfs genes which stimulate mal gene expression with crp^(*1). One the sfs genes (pPVC2) was sequenced and potential CRP binding site is located in the upstream of the putative promoter in the regulatry region. In order to investigate the regulation of the sfs1 gene by the cAMP-CRP complex, we have constructed the sfs-lacZ fusion gene in this research. The overall transcriptional stimulations of sfs1 gene in the presence CAMP were confirmed by β-galactosidase activity and Western blot analysis of sfs1-lacZ fusion gene. Transcriptional regulation by AMP-CRP was also confirmed by Northern blot analysis. End-labelled DNA of the DNA fragment in sfs1 regulatory region were used for gel retardation assay to examine the CRP-DNA complex in the presence of cAMP. Results here indicate that CRP binding site in the regulatory region of sfs1 gene is positive regulator for the expression of sfs1 gene.

Bacillus subtilusHR-1019와 N-Acetyl-thioproline으로 제조한 미생물처리제, BIOACTIVE에 의한 상추의 생장 촉진

Yong-Suk Lee(이용석),Dong-Ju Park(박동주),Jae Hoon Kim(김재훈),Hyeong Seok Kim(김형석),Soo Yeol Chung(정수열),Yong-Lark Choi(최용락) 한국생명과학회 2013 생명과학회지 Vol.23 No.1

L-Cysteine에서 유도된 아미노산인 ATCA와 불용성 인산 가용화능을 가진 균주를 이용하여 미생물처리제 BIOACTIVE를 제조하고 안정성을 확인 하였다. 제조한 미생물처리제를 미생물 비료로 적용하고자 3회 관주 처리한 다음 상추모종의 생장촉진 효과를 조사하는 실험을 수행하였다. 그 결과로 처리한 시험 구에서 상추의 생육은 엽수, 엽장 및 엽중에서 최대 128%, 122% 및 153% 정도로 증가하였다. 처리 간에는 기준량(1,000배 희석) 처리> 반 량(500배 희석) 처리> 배 량(2,000배 희석) 처리 순으로 나타났다. 공시 BIOACTIVE의 처리가 토양 중의 유효인산 및 식물체내의 인산함량을 최대 118% 및 132% 정도 증가 시키는 경향이었다. BIOACTIVE제제를 처리했을 때 N, P, K 3요소를 관행으로 처리와 비슷한 생육 효과가 있는 것으로 보아 복합비료의 사용량을 줄여, 재배가 충분히 가능하리라 생각되며, 상추의 생육 및 비해 조사를 한 결과 본 관주 처리에서는 생육장해 및 비해는 없었다. A biofertilizer, BIOACTIVE, was manufactured by N-acetyl-thioproline (ATCA) and mineral phosphate solubilizing bacteria. The growth promoting effect of the biofertilizer on lettuce was evaluated under three different pot conditions, and its stability was assessed in the field. According to the results of the pot experiments, plant growth was improved compared with that of control: 128%, 122%, and 153% for the leaf number, leaf length, and leaf mass, respectively. Applying the manufactured biofertilizer increased the concentration of phosphate: 118% and 132% in the cultivation soil and plant cells, respectively. These show that BIOACTIVE may have potential as an effective biofertilizer in agriculture.

Klebsiella sp. Sc가 생산하는 β-xylosidase의 분리, 정제 및 특성

Yong-Seok Lee(이용석),In-Hye Park(박인혜),Soon-Cheol Ahn(안순철),Yong-Lark Choi(최용락) 한국생명과학회 2010 생명과학회지 Vol.20 No.12

Klebsiella sp. Sc로부터 birchwood xylan을 분해하는 β-xylosidase를 분리하였다. 이 β-xylosidase는 63 kDa의 분자량을 가지는 559개의 아미노산을 암호화하며 1,680개의 뉴클레오타이드로 구성 되는 것으로 밝혀졌다. 기존에 밝혀진 세균성 β-xylosidase와 상동성을 비교해 보았을 때, Klebsiella oxytoca (KOX)와 90% identities와 95% positives를 나타내었으며 Lactobacillus lactis (LAC, 82%, 90%), Bacillus longum (BLON, 69%, 81%) 그리고 Escherichia coli (ECOLI, 47%, 63%)를 나타내었다. 분리된 β-xylosidase는 GST-fusion 정제 시스템을 이용하여 순수 정제하였다. 이 효소 활성의 최적 pH는 6.6이었으면 최적 온도는 55℃였다. TLC를 통해 효소 분해 산물을 관찰한 결과, xylobiose를 분해하여 xylose를 생산하는 것을 관찰 할 수 있었다. A β-xylosidase encoding gene from Klebsiella sp. Sc was cloned in Escherichia coli. The β-xylosidase gene consisted of an open reading frame of 1,680 nucleotides and encodes 559 amino acids with a deduced molecular weight of 63 kDa. The deduced amino acid sequence of the β-xylosidase from Klebsiella sp. Sc exhibits 90% identities and 95% positives compared to those from Klebsiella oxytoca (KOX), Lactobacillus lactis (LAC, 82%, 90%), Bacillus longum (BLON, 69%, 81%) and Escherichia coli (ECOLI, 47%, 63%). The β-xylosidase was purified by GST-fusion purification system. The pH and temperature optima of the enzyme were 6.6 and 55℃, respectively. The β-xylosidase hydrolyzes xylobiose to xylose.

Isolation and Characterization of a Novel Bacterium, Bacillus subtilis HR-1019, with Insoluble Phosphates Solubilizing Activity

Yong-Suk Lee(이용석),Dong-Ju Park(박동주),Jae Hoon Kim(김재훈),Hyeong Seok Kim(김형석),Yong-Lark Choi(최용락) 한국생명과학회 2013 생명과학회지 Vol.23 No.2

본 연구의 목적은 우수한 미생물 제제 개발용 인산염 가용화 균주의 개발이다. 경작지 토양에서 분리한 인산염 가용화 균주의 특성 및 16S rDNA 염기서열을 조사한 결과 Bacillus subtilis HR-1019로 동정되었다. Bacillus subtilis HR-1019는 hydroxyapatite, tri-calcium phosphate 및 aluminum phosphate 3가지의 난용성 인산염을 모두 가용화하였다. 난용성 인산염의 분해능이 최대가 되는 배양온도는 37℃이었으며, 배양초기 pH가 5.0이었다. 탄소원으로 glucose를 5% 첨가시 가용화능이 높았으며, 가용화된 유리인산의 함량이 증가함에 따라 pH가 크게 감소하였다. 분리균주 HR-1019가 식물병원균에 대하여 균의 생육을 저지하는 clear zone 확인으로 항균효과를 확인하였다. The objective of this study was to develop a mineral phosphate-solubilizing bacterium as a biofertilizer. A mineral phosphate-solubilizing bacterium HR-1019 was isolated from cultivated soils. It was identified as Bacillus subtilis by 16S rDNA analysis. The phosphate-solubilizing activities of the HR-1019 strain against three types of insoluble phosphate, hydroxyapatite, tri-calcium phosphate, and aluminum phosphate were quantitatively determined. When 5% of glucose concentration was used as a carbon source, the strain showed marked mineral phosphate-solubilizing activity. Mineral phosphate solubilization was directly related to pH drop in the culture solution of the strain. The pathogenic activity and antifungal effects of the HR-1019 strain were measured inclear zones formed in PDA media.

Bacillus amyloliiquefaciens AP01 균주에 의한 사이퍼메트린의 생물학적 분해 특성

이용석 ( Yong Suk Lee ),이제훈 ( Je Hoon Lee ),황은정 ( Eun Jung Hwang ),이효정 ( Hyo Jung Lee ),김재훈 ( Jae Hoon Kim ),허재복 ( Jae Bok Heo ),최용락 ( Yong Lark Choi ) 한국응용생명화학회 2016 Journal of Applied Biological Chemistry (J. Appl. Vol.59 No.1

피레스로이드(pyrethroid) 계열 살충제 중 하나인 사이퍼메트린(cypermethrin)의 생물학적 분해를 위하여 부산 인근의 토양과 연안 퇴적물에서 미생물들을 분리하였다. 이들 중 분해율이 우수한 미생물을 선별하여 AP01이라고 명명하였다. 분리 균주 AP01의 유전학적 유연관계를 16S rDNA 염기서열을 분석한 결과, Bacillus amyloliquefaciens와 99%의 상동성을 나타내었다. 다양한 탄소원들 중에서 glucose를 첨가하였을 때, 분리 균주 AP01의 사이퍼메트린(cypermethrin) 분해 활성이 증가하였다. Strain AP01 was isolated for the biological cypermethrin degradation from soil and sediment in Busan. This strain was identified on the basis of phylogenetic analysis of the 16s rDNA sequence and assigned as Bacillus amyloliquefaciens AP01. AP01 could degrade about 45% of cypermethrin in the mineral medium at 30℃ and 180 rpm for 5 days. Furthermore when 2% glucose was added in the medium, the degradation rate of cypermethrin by strain AP01 was increased upto about 60%. Therefore, AP01 may serve as a promising strain in the bioremediation of soil polluted with cypermethrin.

Dissimilatory Nitrate Reduction Characteristics of Indigenous Soybean Rhizobia Distributed in Korea Soils

최영주,최용락,윤한대,유진창,이상규,조무제,Choi, Young-Ju,Choi, Yong-Lark,Yun, Han-Dae,Ryu, Jin-Chang,Lee, Sang-Kyu,Cho, Moo-Je 한국응용생명화학회 1986 Applied Biological Chemistry (Appl Biol Chem) Vol.29 No.2

연속적으로 대두를 재배한 토양과 신개간지 토양에서 재배한 대두 품종 단엽의 근류로부터 토착대두 근류근 87종을 분리하고 이들을 생장속도에 따라 Bradyrhizobium japonicum (slow-grower) 및 Rhizobium fredii (fast-grower)로 분류한 결과 전자가 55종, 후자가 32종이었다. 이두그룹의 대두 근류균을 질산염 환원특성에 따라 denitrifying fast-grower(F-1), nitrate respiring fast-grower(F-II), denitrifying slow-grower(S-I), 및 nitrate respiring slow-grower (S-II)로 세분 되었으며, 분리균주 총 87종중 S-I, S-II, F-I 및 F-II의 수는 각각 10, 22, 48 및 7종이었다. 이들 각 group 대두 근류균의 균체단백질의 전기영동 pattern상의의 차이를 1차 및 2차원 전기영동으로 분석한 결과 slow 및 fast-grower간에는 현저한 차이를 관찰할 수 있었으나 동일 생장속도를 가진 것 중 denitrifer와 nitrate repirer간에는 큰 차이가 없었다. Eightyseven strains of indigenous Rhizobia were isolated from the nodule of soybean cultivar, Danyup, cultivated in four different soils sampled from continuously soybean cultivated and newly reclaimed fields. The strains were grouped into Bradyrhizobium japanicum (slow-grower:55 strains) and Rhizobium fredii (fast-grower: 32 strains). The both groups could be divided into two sub-groups according to the denitrification characteristics, that is, denitrifying fast-grower (F-I), nitrate respiring fast-grower (F-II), denitrifying slow-grower (S-I). and nitrate respiring slow-grower (S-II). Among the 87 isolates, F-I, F-II, S-I and S-II sub-groups were 10, 22, 48 and 7 strains, respectively. The one-and two-dimensional polyacrylamide gel electrophoretic pattern of the four sub-groups were compared and discernible difference was observed between fast and slow-grower, but the difference was not discernible between subgroups within the same growth rate group.

An Effective Method for Isolating Genomic DNA from Leaves of Sesame and Perilla

조영수,최용락,정정한,Cho, Young-Su,Choi, Yong-Lark,Chung, Chung-Han 한국응용생명화학회 1997 Applied Biological Chemistry (Appl Biol Chem) Vol.40 No.3

세포벽 분해 관련 효소의 생산균주 분리 및 CMCase의 특성

이용석(Yong-Seok Lee),박인혜(In-Hye Park),쪼우위(Yi Zhou),김근기(Keun-Ki Kim),최시림(Si-Lim Choi),최용락(Yong-Lark Choi) 한국생명과학회 2008 생명과학회지 Vol.18 No.5

CMCase를 생산하는 Klebsiella sp. Sc를 부산 인근의 토양에서 분리하였다. 이 균주는 16S rDNA와 RNA polymerase B subunit (rpoB) 유전자의 유전적 계통 분석을 통하여 동정을 시행하였다. 그 결과 Klebsiella sp. 속들의 균주들과 99% 이상의 상동성을 나타내었다. CMCase 유전자는 Sc의 게놈라이브러리를 구축하여 찾았다. CMCase 유전자는 37,666 Da의 분자량을 가지는 333개의 아미노산을 암호화하고 있는 1,002개의 뉴클레오티드들로 구성된 것으로 밝혀졌다. 추정된 CMCase의 단백질 서열은 Cellulomonas uda와 단백질과 95%의 identity와 97%의 positive를 가지는 것으로 나타났으며, Erwinia chrysanthemi와는 각각 58%와 72%를 가지는 것으로 나타났다. 그리고 CMCase는 ASDGDTLIAWALLRAQ KQW와 같은 보존 영역을 가지고 있었는데, 이는 glycosyl hydrolase family 8의 보존영역(A-[ST]-D-[AG]-D-X(2)-[IM]-A-[SA]-[LIVM]-[LIVMG]-A-X(3)-[FW])와 일치하는 것으로 나타났다. 이는 Klebsiells sp. Sc의 CMCase는 glycosyl hydrolase family 8에 속한다는 것을 나타낸다. A CMCase-producing bacterium, Klebsiella sp. Sc, was isolated from Busan area's soil sample. This strain was identified on the basis of phylogenetic analysis of the 16S rDNA and rpoB (RNA polymerase B subunit) sequences. The gene encoding CMCase was cloned by genome library. The CMCase gene consisted of an open reading frame of 1,002 nucleotides and encodes 333 amino acids with a deduced molecular weight of 37,666 Da. The deduced amino acid sequence of the CMCase from Klebsiella sp. exhibits 95% identities and 97% positives to those from Cellulomonas uda and 58% identities and 72% positives to those from Erwinia chrysanthemi. The CMCase contain a conserved motif with the consensus sequence, ASDGDTLI AWALLRAQKQW. This result shows CMCase protein from Klebsiella sp. Sc belongs to glycoside hydrolase family 8, A-[ST]-D-[AG]-D-X(2)-[IM]-A-[SA]-[LIVM]-[LIVMG]-A-X(3)-[FW].

Molecular Cloning of Alfalfa(Medicago Sativa L. Vernal) Leghemoglobin Structural Genes Synthesized by $Poly(A)^+$-mRNA from Alfalfa Root Nodules

조무제,윤한대,최용락,최영주,이경상,Cho, Moo-Je,Yun, Han-Dae,Choi, Yong-Lark,Choi, Young-Ju,Lee, Kyung-Sang,Brill, Winston J. 생화학분자생물학회 1985 한국생화학회지 Vol.18 No.1

알팔파 근류에서 분리 정제한 $poly(A)^+$-mRNA로부터 leghemoglobin message 함유비율이 높은 9S, 18S 및 28S mRNA를 이용하여 cDNA를 합성하였다. 이들 cDNA를 Sal I linker에 연결 pBR322의 Sal I site에 크로닝하여 cDNA library를 만든 후 이들 library로부터 colony hybridization, Southern blot hybridization, hybrid-released 및 hybrid-arrestes translation에 의하여 leghemoglobin cDNA clone을 동정하였으며, 동정된 clone으로부터 alfalfa leghemoglobin cDNA의 제한효소 지도를 작성하였다. Complementary DNAs were synthesized by the leghemoglobin message rich 9S, 18S and 28S-mRNA fractions prepared from the $poly(A)^+$-mRNA of alfalfa root nodules. The synthetic Sal I linkered double stranded cDNA was inserted into the Sal I site of the pBR322, and transformed in E. coli HB101. The leghemoglobin cDNA clones were screened by colony hybridization and Southern blot hybridization with the soybean leghemoglobin cDNA as a probe, and further screened by hybrid-released and hybrid-arrested translation. One alfalfa leghemoglobin cDNA clone was selected, and restriction map and fingerprints of the cDNA from the selected clone were analyzed.

알파파 근류로부터 분리한 Poly ( A+ ) - mRNA 에 의하여 합성된 Leghemoglobin 유전자의 cDNA Cloning 에 관한연구

조무제,윤한대,최용락,최영주,이경상,Winstion J . Brill ( Moo Je Cho,Min Suk Yang,Han Dae Yun,Yong Lark Choi,Young Ju Choi,Kyung Sang Lee ) 생화학분자생물학회 1985 BMB Reports Vol.18 No.1

Complementary DNAs were synthesized by the leghemoglobin message rich 9S, 18S and 28S-mRNA fractions prepared from the poly(A)^+-mRNA of alfalfa root nodules. The synthetic Sal I linkered double stranded cDNA was inserted into the SalI site of the pBR322, and transformed in E. coli HB101. The leghemoglobin cDNA clones were screened by colony hybridization and Southern blot hybridization with the soybean leghemoglobin cDNA as a probe, and further screened by hybrid-released and hybridarrested translation. One alfalfa leghemoglobin cDNA clone was selected, and restriction map and fingerprints of the cDNA from the selected clone were analyzed.