• Microbiology and Biotechnology Letters
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• v.17 no.3
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• pp.178-182
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• 1989

A gene coding for a xylanase of thermophilic alkalophilic Bacillus sp. K-17 was cloned in Escherichia coli C600 with pBR322. Plasmid pAXl13 was isolated from a transformant producing xylanase, and the xylanase gene was located in a 4.3 Kb HindIII fragment. Biotinylated pAXl13 hybridized to a 4.3 Kb HindIII fragment from chromosomal DNA of thermophilic alkalophilic Bacillus sp. K-17. The xylanase activity was observed in the extracellular curture fluid of E. coli carrying pAXl13. The pAXl13-encoded xylanase had the same enzymatic properties as those of xylanase I produced by thermophilic alkalophilic Bacillus sp. K-17.

• The Microorganisms and Industry
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• v.20 no.3
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• pp.2-13
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• 1994

PHA 생합성에 관련된 기초 연구도 미생물학, 생화학, 그리고 분자생물학 각도에서 활발히 수행되어, 신규 PHA 생합성 미생물의 탐색, 대사경로 및 조절 mechanism의 규명, 그리고 물성이 개량된 각종 PHA 공중합체의 개발 연구가 활발히 이루어졌다. 특히 최근에는 recombinant DNA 기술을 이용한 각종 미생물 유래의 PHA 생합성 관련 유전자의 분리와 그 기능에 관한 연구가 활발히 수행되고 있고, 1988년 A. eutrophus의 PHB 생합성 관련 세개의 효소를 coding하는 유전자가 독일의 Steinbchel등(5), 미국의 Dennis 등(6), 그리고 Sinskey등(7,8)에 의해 거의 동시에 cloning되었으며, 이를 이용한 대사 경로 및 조절 기작에 관한 연구가 본격화 되었다. 또한 최근에는 재조합 균주를 이용한 PHA의 생산에 관한 연구도 활발히 진행되고 있으며, 이와 같은 최근의 연구 성과는 몇 편의 총설(9-12)에 잘 요약되어 있다.

• Journal of Life Science
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• v.27 no.12
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• pp.1403-1409
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• 2017

In this study, the xylitol dehydrogenase (XYL2) gene was expressed in Saccharomyces cerevisiae as a host cell for ease of use in the degradation of lignocellulosic biomass (xylose). To select suitable expression systems for the S.XYL2 gene from S. cerevisiae and the P.XYL2 gene from Pichia stipitis, $pGMF{\alpha}-S.XYL2$, $pGMF{\alpha}-P.XYL2$, $pAMF{\alpha}-S.XYL2$ and $pAMF{\alpha}-P.XYL2$ plasmids with the GAL10 promoter and ADH1 promoter, respectively, were constructed. The mating factor ${\alpha}$ ($MF{\alpha}$) signal sequence was also connected to each promoter to allow secretion. Each plasmid was transformed into S. cerevisiae $SEY2102{\Delta}trp1$ strain and the xylitol dehydrogenase activity was investigated. The GAL10 promoter proved more suitable than the ADH1 promoter for expression of the XYL2 gene, and the xylitol dehydrogenase activity from P. stipitis was twice that from S. cerevisiae. The xylitol dehydrogenase showed $NAD^+$-dependent activity and about 77% of the recombinant xylitol dehydrogenase was secreted into the periplasmic space of the $SEY2102{\Delta}trp1/pGMF{\alpha}-P.XYL2$ strain. The xylitol dehydrogenase activity was increased by up to 41% when a glucose/xylose mixture was supplied as a carbon source, rather than glucose alone. The expression system and culture conditions optimized in this study resulted in large amounts of xylitol dehydrogenase using S. cerevisiae as the host strain, indicating the potential of this expression system for use in bioethanol production and industrial applications.

• Korean Journal of Microbiology
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• v.30 no.1
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• pp.1-7
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• 1992

We cloned a gene cluster encoding phenanthrene-degrading enzymes on a 6.8-kb Xhol fragment from the Pseudomonas sp. DJ77 chromosomal DNA into the vector pBLUESCRIPT SIC(+). The resultant clone, containing the recombinant plilsmid pHENX7, was able to convert 3-methylcatechol to a yellow mela-cleavage compound. Since the pHENX7R in which the DNA insert was cloned in the opposite orientation lacked extradiol dioxygenase activity. the direction of transcription was established. Four polypeptides, PhnC (24 kDa). PhnD (31 kDa), PhnE (34 kDa). and PhnF (15 kDa), were identified in E coli JM101 transformed with several pHENX7-derived plasmids. The locations and extents of ~ndividual genes were determined by subcloning. The gene order was phnC-phnD-phnE-phnF-phnG, and phnC, phnD, phnE, and phnG genes encoded glutathione S-transferase, mrta-cleavage compound hydrolase, extradiol dioxygenase, mera-cleavage compound dehydrogenase, respectively.

• Korean Journal of Microbiology
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• v.29 no.6
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• pp.392-396
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• 1991

Mu d1(Ap lac) bacteriophage can be used to search for genes which are members of a common regulatory network without having to know the functions of the genes in advance. Aim was for obtaining the loci in the SOS network as well as temperature inducible loci. For this purpose, recA441 allele was used. This allele encodes a thermosensitive recA gene product; thus, the recA441 allele can be activated upon temperature upshift without by external DNA damage. Approximately 10, 000 colonies were screened, and then searched for the colonies which expressed .betha.-galactosidase higher level at 42.deg.C than at 30.deg.C. The strains identified fell into two dlasses; (i) ones in which the increased expression was $recA^{+}$ $lexA^{+}$ -dependent, that is, din(damage-inducible) genes which were due to the activation of recA441 allele and (ii) ones in which the increased expression was $recA^{+}$ $lexA^{+}$ -independent and only temperature-inducible, tin genes. Rough mapping position was obtained for these genes.

• KSBB Journal
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• v.22 no.1
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• pp.62-65
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• 2007

New strain development strategy using kinetic models and metabolic control analysis was investigated. In this study, previously reported mathematical models describing the enzyme kinetics of intracellular threonine synthesis were modified for mutant threonine producer Escherichia coli TF5015. Using the modified models, metabolic control analysis was carried out to identify the rate limiting step by evaluating the flux control coefficient on the overall threonine synthesis flux exerted by individual enzymatic reactions. The result suggested the production of threonine could be enhanced most efficiently by increasing aspartate semialdehyde dehydrogenase (asd) activity of this strain. Amplification of asd gene in recombinant strain TF5015 (pCL-$P_{aroF}$-asd) increased the threonine production up to 23%, which is much higher than 14% obtained by amplifying aspartate kinse (thrA), other gene in threonine biosynthesis pathway.

• Journal of Life Science
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• v.18 no.1
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• pp.136-142
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• 2008

A recombinant Pichia pastoris carrying double expression cassette of Rhodotorula glutinis epoxide hydrolase(RgEH) gene was developed and used for preparing enantiopure (S)-styrene oxide from racemic mixture of styrene oxide. BglII restriction site of original RgEH gene (pPICZ B/RgEH #2) of previous report was mutated using PCR technique for the construction of double expression cassette containing promoter ($P_{AOX1}$), EH gene and transcription terminator ($TT_{AOX1}$) in pPICZ C vector. Double expression cassette with RgEH was inserted into the chromosomal DNA of P. pastoris. $V_{max}$ ($2.2{\mu}mol\;min^{-1}mg\;dcw^{-1}$) on (R)-styrene oxide of P. pastoris with double expression cassette was about 6-fold higher than that ($0.4{\mu}mol\;min^{-1}mg\;dcw^{-1}$) of P. pastoris with single expression cassette. For the determination of the optimal condition, the effects of detergent and temperature on the enantioselective hydrolytic activity and yield of the enantiomer were investigated. When the reaction was performed at $10^{\circ}C$ for 10 min in the presence of 0.5% Tween 20, enantiopure (S)-styrene oxide with 99.9% ee was obtained as the yield 43.4 % from 20 mM racemic sustrate.

• KSBB Journal
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• v.21 no.5
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• pp.394-400
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• 2006

When the papain, which is a sort of Cystein protease, is applied to the outer skin, it decomposes the protein which forms the peeled outer skin and speeds up metabolism. Therefore, it is one of the most important cosmetics compositic which keeps the function of skin normal. When the papain is used in cosmetics with surfactant, the activity of papain is reduced rapidly. In this study, the modified papain with extreme stability negative ionic environment was developed by directed evolution

• Korean Journal of Microbiology
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• v.32 no.2
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• pp.126-131
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• 1994

The pchABCD genes in Pseudomonas sp. DJ-12 speciyin degradation o 4-chlorobiphenyl(4CB) were cloned in Eschericia coli. The cloned cells of E. coli CU1 and CU101 showed to produce 2,3-dihydroxybiphenyl (2,3-DHBP) from 4-chlorobiphenyl by dechlorination, as Pseudomonas so. DJ-12 produced 2,3-DHBP from both biphenyl and 4CB. In particular, E. coli CU101 transformed with the recombinant plasmid of pCU101 revealed dechlorination activity to produce 2,3-DHBP from 4CB without production of 4-chlorobenzoic acid. Therefore, the pcbAB genes (2.2 kb in size) cloned from the chromosome of Pseudomonas sp. DJ-12 were found to have dechlorination activity on 4CB to produce 2,3-DHNP.

• Journal of Food Hygiene and Safety
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• v.6 no.3
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• pp.133-137
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• 1991

In order to increase the expression of XMP aminase [EC 6.3.4.1], which catalizes the conversion of 5'-XMP to the DNA fragment containing gua A gene coding for XMP aminase from pLC 34-10 plasmid was subcloned into pBR 322, and 1.7 kb gua A gene fragment was recloned under the control of trp promoter of pDR 720, E. coli expression vector. XMP aminase activity had increased by about 17 times when compared with that of the strain earring pLC 34-10.