• Journal of Microbiology and Biotechnology
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• 제13권1호
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• pp.90-98
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• 2003

In vivo $^31p$ Nuclear Magnetic Resonance ($^31p$NMR) and metabolic studies were carried out on an acetic acid tolerant mutant, Zymomonas mobilis $ZM4/Ac^R$, and compared to those of the parent strain, Z. mobilis ZM4, to evaluate possible mechanisms of acetic acid resistance. This investigation was initiated to determine whether or not the mutant strain might be used as a suitable recombinant host far ethanol production from lignocellulose hydrolysates containing various inhibitory compounds. $ZM4/Ac^R$ showed multiple resistance to other lignocellulosic toxic compounds such as syringaldehyde, furfural, hydroxymethyl furfural, vanillin, and vanillic acid. The mutant strain was resistant to higher concentrations of ethanol or lower pH in the presence of sodium acetate, compared to ZM4 which showed more additive inhibition. in vivo $^31p$ NMR studies revealed that intracellular acidification and de-energization were two mechanisms by which acetic acid exerted its inhibitory effect. For $ZM4/Ac^R$, the internal pH and the energy status were less affected by sodium acetate compared to the parent strain. This resistance to pH change and de-energization caused by acetic acid is a possible explanation for the development of resistance by this strain.

• 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.

• 한국미생물·생명공학회지
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• 제18권3호
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• pp.268-272
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• 1990

Zymomonas mobilis의 알코올 탈수소 효소 유전자가 클로닝된 대장균 형질전환체의 세포 추출물로부터 알코올 탈수소 효소를 분리정제하였다. 형질전환된 Escherichia coli(pADS93)가 생산하는 Z.mobilis 유전자 유래의 알코올 탈수소 효소는 분자량이 40,000인 동일한 4개의 subunits로 구성된 tetramer임이 밝혀졌으며 이것은 Z.mobilis의 세포 추출물로부터 분리한 알코올 탈수소 효소와 동일하였다. 이 효소의 정반응(ethanol 산화)은 pH의 영향을 많이 받으며 pH는 10.0이었고 역반응(acetaldehyde 환원)에서는 최적의 pH가 7.5-8.5 이었지만 pH에 따라 크게 영향을 받지는 않았다.

• 한국미생물·생명공학회지
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• 제17권6호
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• pp.600-605
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• 1989

Escherichia coli 내에서 프로모터활성을 보이는 Zymomonas mobilis 유래의 유전자 절편을 분리하고 특성을 분석하였다. 프로모터 탐색용 벡터인 pCMT215는 promoter activity가 없는 pMT21의 HinIII 위치에 pYEJ001의 클로람페니콜 아세틸전이 효소유전자를 함유한 0.7-kb HindIII 조각을 접합시켜 제조하였다. E. mobilis의 chromosomal DNA를 Sau3AI으로 부분절단하여 pCMT215에 도입한 후, 이를 이용하여 대장균을 형질전환시킨 결과 14개의 형질전환주가 선별되었다. 이들은 30-750 $\mu$g/$m\ell$ 농도의 chloramphenicol에 내성을 보였으며 클로닝된 유전자조각의 크기는 0.1-1.5Kb였다. 이 가운데 5개의 염기서열을 분석해 본 결과 일반적인 프로모터의 염기서열과 많은 유사점이 발견되었는데, 대장균의 프로모터인 -35 또는 -10 지역과의 부분적인 일치와 A 또는 T 염기가 풍부한 지역과 연속적인 A 또는 T 염기배열, 그리고 회문형태의 염기서열 등이 발견되었다. 또한 대장균 내에서의 프라이머 연장실험결과 Z. mobilis로부터 유래된 DNA조각에서 전사의 시작이 4-170 염기의 거리를 두고 두 곳 또는 여러 곳에서 일어남을 알 수 있었다.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 1986년도 추계학술대회
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• pp.521.3-522
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• 1986

A gene encoding alcohol dehydrogenase (ADH) in Zymomonas mobilis was cloned into E. coli JM 83 with plasmid pUC 9. The ADH produced by the E. coli transformant was purified bysonication, (NH$^4$)2SO4 fractionation, Affi-Gel blue and hydroxylapatite chromatography. The ADH produced by Z. mobilis was also purified by the same procedures. The two enzyme preparations were characterized and compared. It was found that the E. coli ADH was identical to one of two ADH isozymes of Z. mobilis. Analytical gel filtrations led to the conclusion that the molecule of E. coli ADH was composedv of four subunits having molecular weight of 40,000 (+1,000) dalton each The effect of metal ions on ADH activity and optimum pH were investigated.

• Journal of Microbiology and Biotechnology
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• 제1권1호
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• pp.12-16
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• 1991

Previously RP1::Tn951 which is a derivative of RP1 containing the lactose transposon Tn951 was introduced into Z. mobilis strain ZM6l00, and RP1::Tn951 was integrated into its genome to yield ZM6306. The galactose operon was incorporated into ZM6306 to yield ZM6307 for more efficient utilization of lactose. Batch culture study has been carried out on Z. mobilis strains, ZM6306 ($lac^+$ ) and ZM6307 ($lac^+$ , $gal^+$ ), which can convert lactose directly to ethanol. Using a medium containing 80 gㆍ$1^{-1}$ glucose and 40 gㆍ$1^{-1}$ lactose, it was found that ZM6306 and ZM6307 produced maximum ethanol concentration of 40 gㆍ$1^{-1}$ and 42 gㆍ$1^{-1}$, respectively, whereas parent strain ZM6 produced 37 gㆍ$1^{-1}$.

• KSBB Journal
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• 제11권1호
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• pp.58-64
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• 1996

K-carrageenan에 고정화시킨 Zymomonas mobilis를 이용한 연속적인 sorbitol 생산에 대하여 연구하였다. Toluene과 glutaraldehyde로 처리하여 투과성을 증대시킨 세포를 alginate나 chitin 고정화 공정에서 어느 정도의 효소활성을 보였으나 210시간 이상의 공정반응에서는 sorbitol 생성량이 감소되었다. 독성이 적고 투과성을 증가시키는 물질로서 toluene 대신 CTAB(Cetyltrimethylammoniumbromide)를 사용하였다. CTAB로 투과성을 증가시킨 세포를 k-carrageenan에 고정화하여 CSTR과 packed bed reactor를 이용하여 sorbitol 생산을 시도하였으며 CSTR보다도 (25일) packed bed reactor에서 (30일) 더 긴 시간 효소활성도가 유지되었다. Two-stage 연속공정에서는 희석비율이 (dilution rate, $h^{-1}$) 증가함에 따라서 sorbitol 생성률이 증가되었으며 희석비율 0.32 $h^{-1}$에서 첫번째와 두번째 반응조에서 각각 $15g/\ell-h,$ $22g/\ell-h의$ sorbitol 생성률을 얻었다. 0.32$h^{-1}$보다 높은 희석비율에서는 생성를이 감소되었다.

• KSBB Journal
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• 제7권1호
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• pp.15-20
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• 1992

식품첨가물 등으로 이용 되어지는 sorbitol을 $\beta$-1,2fructose oligomer를 함유하고 있는 Jerusalem artichoke를 이용하여 생산 하였으며, 이는 L-sorbose생산에 있어서 전구물질로 이용된다. Inulinase와 oxidoreductase의 동시고정화여, inulin을 와 fructose로 가수분해함과 동시에 가수분해산물을 sorbitol로 전환시켰다. Inulinase와 oxidoreductase는 chitin(5%, w/v)과 K-carrageenan(4%, w/v)으로 고정화 시켰다. 0.2% CTAB(Cetyltrimethylammonlumbromide)처리에 의해서 투과성이 향상된 Zymomonas mobilis세포내의 함유 되어있는 oxidoreductase의 activity가 향상되었다. Jerusalem artichoke juice를 acid에 의한 가수분해에 의해서 40%(v/w) juice 100ml당 53.64의 total carbohydrate yield를 얻었고, 온도 변화에 따른 가수분해시 $85^{\circ}C$에서 반응이 신속히 일어났다. Inulinase를 이용한 가수분해에 의하여 35.56g/l의 glucose와 85.32g/l의 fructose가 생성되었으며, sorbitol생성에 기질로 이용되었다. Inulinase와 Z. mobilis세포의 동시고정화 반응기에서 35.15g/l의 sorbitol을 생성하였으며, 이는 동시고정화를 하지 않았을때의 sorbitol생성농도(35.64g/l)보다 낮았다.

• Journal of Microbiology and Biotechnology
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• 제6권1호
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• pp.36-42
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• 1996

The production of sorbitol by permeabilized cells of Zymomonas mobilis immobilized in $\kappa$-carrageenan was investigated. Cetyltrimethylammoniumbromide (CTAB) permeabilized cells were treated with glutaraldehyde prior to immobilization for cross-linking of enzymes, glucose-fructose oxidoreductase (GFOR) in cells. Rigidity of the immobilized beads was increased two-fold with 90$\%$ conversion efficiency by the additions of 40$\%$ (w/v) polyols (glycerol 25 g + propylene glycol 15 g) to 60$\%$ (w/v) distilled water containing 2.5$\%$ (w/v) $\kappa$-carrageenan as a final concentration, prior to immobilization. $\kappa$-Carrageenan beads entrapping permeabilized cells were dried to improve bead rigidity and storage stability. During s6mi-batch process for 72 h with dry beads, there was an improvement of the loss of enzyme activity (less than 10$\%$). In batch process, the kinetic results of $K_m.fructose$ value for the free cells, wet beads and dry $\kappa$-carrageenan beads were 71.7, 72.4 and 116.7 g/l, respectively. Higher productivity was obtained with two-stage continuous packed bed reactors with both wet and dry $\kappa$-carrageenan beads at 25.00 and 21.15 g/l/h, respectively, when measured at second stage.

• Mycobiology
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• 제32권4호
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• pp.170-172
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• 2004

The efficiency of acid, enzyme and microbial pretreatment of rice bran was compared based on the content of cellulose, hemicellulose, reducing sugars and xylose in the substrate. An isolate of Aspergillus niger or a strain of Trichoderma viride(MTCC 800) was employed for microbial pretreatment of rice bran in solid state. Acid pretreatment resulted in the highest amount of reducing sugars followed by enzyme and microbial pretreatment. A. niger showed a higher rate of hydrolysis than T. viride. The rice bran hydrolysate obtained from the different methods was subsequently fermented to ethanol either by Zymomonas mobilis(NCIM 806) or by Pichia stipitis(NCIM 3497). P. stipitis fermentation resulted in higher ethanol(37% higher) and biomass production($76{\sim}83%$ higher) than those of Z. mobilis. Maximum ethanol production resulted at 12h in Zymomonas fermentation, while in Pichia fermentation, it was observed at 60h. Microbial pretreatment of rice bran by A. niger followed by fermentation employing P. stipitis was more efficient but slower than the other microbial pretreatment and fermentation.