• Journal of Microbiology and Biotechnology
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• 제23권2호
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• pp.251-259
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• 2013

An endoxylanase gene (PoxynA) that belongs to the glycoside hydrolase (GH) family 11 was cloned from a xylanolytic strain, Penicillium oxalicum B3-11(2). PoxynA was overexpressed in Trichoderma reesei QM9414 by using a constitutive strong promoter of the encoding pyruvate decarboxylase (pdc). The high extracellular xylanase activities in the fermentation liquid of the transformants were maintained 29~35-fold higher compared with the wild strain. The recombinant POXYNA was purified to homogeneity, and its characters were analyzed. Its optimal temperature and pH value were $50^{\circ}C$ and 5.0, respectively. The enzyme was stable at a pH range of 2.0 to 7.0. Using beechwood as the substrate, POXYNA had a high specific activity of $1,856{\pm}53.5$ IU/mg. In the presence of metal ions, such as $Cu^{2+}$, and $Mg^{2+}$, the activity of the enzyme increased. However, strong inhibition of the enzyme activity was observed in the presence of $Mn^{2+}$ and $Fe^{2+}$. The recombinant POXYNA hydrolyzed birchwood xylan, beechwood xylan, and oat spelt xylan to produce short-chain xylooligosaccharides, xylopentaose, xylotriose, and xylobiose as the main products. This is the first report on the expression properties of a recombinant endoxylanase gene from Penicillium oxalicum. The properties of this endoxylanase make it promising for applications in the food and feed industries.

• 한국식품저장유통학회지
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• 제18권5호
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• pp.795-802
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• 2011

Carbonic maceration처리 포도주에 있어 유기산 특히 사과산 함량을 감소시키는 주요 원인을 찾고자 포도를 2주 동안 온도별 carbonic maceration 처리하며 시기별로 산함량 및 사과산대사 관련 효소활성을 측정하였다. 온도별 carbonic maceration 처리 포도의 pH는 CM-$25^{\circ}C$와 CM-$35^{\circ}C$에서 처리 시간이 경과할수록 가장 높았으며, 총산은 초기에는 CM-$35^{\circ}C$에서 가장 낮은 함량을 나타내었지만 처리 6일 이후 서서히 증가하였고, CM-$25^{\circ}C$는 처리 완료일 까지 꾸준히 감소하는 것으로 나타났다. CM-$45^{\circ}C$는 초기와 비슷한 함량을 나타내어 다른 처리보다 높은 총산 함량을 나타내었다. 유기산 함량에 있어 사과산 함량은 CM-$35^{\circ}C$에서 가장 많이 감소하였고, 젖산 함량은 CM-$35^{\circ}C$에서 가장 높게 나타났다. 사과산 대사과련 효소활성을 살펴본 결과, malic enzyme과 malic dehydrogenase는 CM-$25^{\circ}C$와 CM-$35^{\circ}C$에서 가장 높은 효소활성을 나타내었지만 CM-$45^{\circ}C$에서는 초기부터 효소활성이 나타나지 않았다. oxalacetate decarboxylase 활성도 malic dehydrogenase 활성과 비슷한 경향을 나타내었다. pyruvate decarboxylase 활성은 다른 효소활성에 비해 활성이 낮았지만, CM-$45^{\circ}C$에서도 활성을 나타내었다. 반면 L-lactic dehydrogenase 활성은 어떤 처리구에서도 나타나지 않았다. 이와 같은 결과로부터 온도와 효소활성과의 관계에 있어 온도가 $40^{\circ}C$ 이상에서는 사과산 대사관련 효소가 활성을 나타내지 않는 것을 알 수 있었고, carbonic maceration 처리에서 사과산 감소가 효소의 작용에 크게 영향 받는 것을 확인할 수 있었지만, 젖산 생성에 대해서는 효소작용 외에 사과산 대사 미생물과 같은 다른 요인들에 대해 좀 더 깊이 있는 연구가 필요하다.

• Journal of Microbiology and Biotechnology
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• 제21권10호
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• pp.1049-1052
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• 2011

In this study, we demonstrated the asymmetric synthesis of L-tert-leucine from trimethylpyruvate using branched-chain aminotransferase (BCAT) from Escherichia coli in the presence of L-glutamate as an amino donor. Since BCAT was severely inhibited by 2-ketoglutarate, in order to overcome this here, we developed a BCAT/aspartate aminotransferase (AspAT) and BCAT/AspAT/pyruvate decarboxylase (PDC) coupling reaction. In the BCAT/AspAT/PDC coupling reaction, 89.2 mM L-tert-leucine (ee>99%) was asymmetrically synthesized from 100 mM trimethylpyruvate.

• Journal of Microbiology and Biotechnology
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• 제13권4호
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• pp.571-577
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• 2003

The intracellular metabolic fluxes can be calculated by metabolic flux analysis, which uses a stoichiometric model for the intracellulal reactions along with mass balances around the intracellular metabolites. In this study, metabolic flux analyses were carried out to estimate flux distributions for the maximum in silico yields of various metabolites in Escherichia coli. The maximum in silico yields of acetic acid and lactic acid were identical to their theoretical yields. On the other hand, the in silico yields of succinic acid and ethanol were only 83% and 6.5% of their theoretical yields, respectively. The lower in silico yield of succinic acid was found to be due to the insufficient reducing power. but this lower yield could be increased to its theoretical yield by supplying more reducing power. The maximum theoretical yield of ethanol could be achieved, when a reaction catalyzed by pyruvate decarboxylase was added in the metabolic network. Futhermore, optimal metabolic pathways for the production of various metabolites could be proposed, based on the results of metabolic flux analyses. In the case of succinic acid production, it was found that the pyruvate carboxylation pathway should be used for its optimal production in E. coli rather than the phosphoenolpyruvate carboxylation pathway.

• Journal of Microbiology and Biotechnology
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• 제20권7호
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• pp.1156-1162
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• 2010

During ethanol fermentation, bacterial strains may encounter various stresses, such as ethanol and acid shock, which adversely affect cell viability and the production of ethanol. Therefore, ethanologenic strains that tolerate abiotic stresses are highly desirable. Bacteria of the genus Deinococcus are extremely resistant to ionizing radiation, ultraviolet light, and desiccation, and therefore constitute an important pool of extreme resistance genes. The irrE gene encodes a general switch responsible for the extreme radioresistance of D. radiodurans. Here, we present evidence that IrrE, acting as a global regulator, confers high stress tolerance to a Zymomonas mobilis strain. Expression of the gene protected Z. mobilis cells against ethanol, acid, osmotic, and thermal shocks. It also markedly improved cell viability, the expression levels and enzyme activities of pyruvate decarboxylase and alcohol dehydrogenase, and the production of ethanol under both ethanol and acid stresses. These data suggest that irrE is a potentially promising gene for improving the abiotic stress tolerance of ethanologenic bacterial strains.

• 한국작물학회지
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• 제52권3호
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• pp.281-288
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• 2007

Photoperiod sensitive genetic male sterile (PGMS) rice is sterile mutant controlled by photoperiod. A PGMS mutant 920S was sterile grown under long-day (LD) photoperiod (14 h light/10 h dark) but fertile grown under short-day (SD) photoperiod (10 h light/14 h dark). Proteome analysis revealed that 12 protein spots were differentially expressed in the spikelets of 920S plants either treated with LD or SD photoperiod. Among these proteins, three proteins including chlorophyll a/b binding protein, vacuolar ATPase ${\beta}-subunit,\;{\alpha}-tubulin$ and an unknown protein were more than three-fold abundant in the spikelet of the SD-treated plants than those of the LD-treated plants. On the other hand, eight proteins including acetyl transferase, 2, 3- biphosphoglycerate, aminopeptidase N, pyruvate decarboxylase, 60S acidic ribosomal protein and three unknown protein spots were more abundant in the spikelets of the LD-treated plants than those of the SD-treated plants. The results suggest that the observed proteins may be involved in sterile or fertile pollen development under LD or SD photoperiod respectively in the PGMS mutant rice.

• Molecules and Cells
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• 제24권3호
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• pp.394-408
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• 2007

Several genes/QTLs governing resistance/tolerance to abiotic and biotic stresses have been reported and mapped in rice. A QTL for submergence tolerance was found to be co-located with a major QTL for broad-spectrum bacterial leaf blight (bs-blb) resistance on the long arm of chromosome 5 in indica cultivars FR13A and IET8585. Using the Nipponbare (japonica) and 93-11 (indica) genome sequences, we identified, in silico, candidate genes in the chromosomal region [Kottapalli et al. (2006)]. Transcriptional profiling of FR13A and IET8585 using a rice 22K oligo array validated the above findings. Based on in silico analysis and arraying we observed that both cultivars respond to the above stresses through a common signaling system involving protein kinases, adenosine mono phosphate kinase, leucine rich repeat, PDZ/DHR/GLGF, and response regulator receiver protein. The combined approaches suggest that transcription factor EREBP on long arm of chromosome 5 regulates both submergence tolerance and blb resistance. Pyruvate decarboxylase and alcohol dehydrogenase, co-located in the same region, are candidate downstream genes for submergence tolerance at the seedling stage, and t-snare for bs-blb resistance. We also detected up-regulation of novel defense/stress-related genes including those encoding fumaryl aceto acetate (FAA) hydrolase, scramblase, and galactose oxidase, in response to the imposed stresses.

• 한국작물학회:학술대회논문집
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• 한국작물학회 2017년도 9th Asian Crop Science Association conference
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• pp.81-81
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• 2017

The origin of wild corn (teosinte) is distributed in the Northwest coastal pacific area of Central America, including Mexico, which is a wetland area of 5 to 6 months per year. Depending on these climate characteristics, wild corn is genetically resistant to flooding condition. In order to evaluate the availability of flooding resistant genes of these wild corns, we examined the physiological responses after the flooding treatment in the early stages of the growth of various wild corns. The difference of chlorophyll content between flooding untreatment and flooding treatments (untreated chlorophyll content - humidified chlorophyll content) was the highest in chlorophyll content in the case of B73, the common corn. In the middle leaf, $\underline{Zea}$ mays subsp. Parviglumis, Zea mays subsp. Mexicana, Zea mays subsp., Zea perennis decreased significantly. In the lower leaves, Zea mays sub and Zea nicaraguensis showed the lowest content compared to B73. PCR analysis was performed using 34 primers divided into two groups, top and bottom. In the wild corn, pyruvate decarboxylase 2 in root and alcohol dehydrogenase 1 in shoot showed the difference in the reaction.

• Journal of Plant Biotechnology
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• 제4권1호
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• pp.7-15
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• 2002

Eight cDNAs corresponding to fruit-specific genes were isolated from ripened melon through differential screening. Sequence comparison indicated that six of these cDNAs encoded proteins were previously characterized into aminocyclopropane-1-carboxylate (ACC) oxidase, abscisic acid, stress and ripening inducible (ASR) gene, RINC-H2 zinc finger protein, pyruvate decarboxylase, or polyubiquitin. RFS2 and RFS5 were the same clone encoding polyubiquitin. The other cDNAs showed no significant homology with known protein sequences. The ASR homologue (Asr1) gene was further characterized on the cDNA and genomic structure. The deduced amino acid sequence had similar characteristics to other plant ASR. The Asr1 genomic DNA consisted of 2 exons and 1 intron, which is similar to the structure of other plants ASR genes. The promoter region of the Asr1 gene contained several putative functional cis-elements such as an abscisic acid responsive element (ABRE), an ethylene responsive element (ERE), a C-box or DPBf-1 and 2, Myb binding sites, a low temperature responsive element (LTRE) and a metal responsive element (MRE). The findings imply that these elements may play important roles in the response to plant hormones and environmental stresses in the process of fruit development. The results of this study suggest that the expressions of fruit specific and ripening-related cDNAs are closely associated with the stress response.

• 한국미생물·생명공학회지
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• 제35권4호
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• pp.272-277
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• 2007

P. pastoris는 대장균에 비해 정확한 접힘, 당화, 효율적인 분비기작등의 장점을 가지고 있어 재조합 단백질의 생산을 위한 균주로서 관심을 받고 있다. 또한 재조합 단백질의 효율적인 생산공정 개발을 위하여 배양공정 중 특정 시간이나 조건에서 발현될 수 있는 효율적인 유도성 프로모터의 개발도 중요 관심 분야이다. 본 연구에서는 연속배양을 이용하여 P. pastoris의 배양 중 특정 기질이 소모되었을 때 발현되는 자동 유도성 프로모터를 개발하기 위하여, 인산이 고갈되었을 때 과발현 되는 유전자들을 탐색하였다. 인산 제한 연속배양의 정상상태에서 얻어진 균체로 부터 total RNA와 mRNA를 분리하였고, 이로부터 cDN를 합성하여 인산제한조건에서 과발현되는 유전자들을 확보하였다. 그 중 빈도수가 높은 8종의 유전자 3-phosphoglycerate kinase, glyceraldehyde 3-phosphate dehydrogenase(GAPDH), glucokinase, thiol-specific antioxidant protein, triosephosphate isomerase, sodium/phosphate symporter(NPS) 그리고 pyruvate decarboxylase를 선별하였고, Northern blot analysis를 수행한 결과 인산 섭취에 관련된 NPS 유전자가 인산제한조건에서 과발현 됨을 확인하였다. 본 연구실에서는 자동유도성 프로모터로서 NPS유래의 프로모터의 잠재성을 알아보기 위하여, 관련 유전자를 확보하여 외래 유전자를 이용한 발현연구를 진행 중이다.