• Journal of Microbiology and Biotechnology
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• 제12권3호
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• pp.411-416
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• 2002

Bacillus macerans cyclodextrin glucanotransferase (CGTase) was expressed on the cell surface of Saccharomyces cerevisiae by fusing with Aga2p linked to the membrane-anchored protein, Aga1p. The surface display of CGTase was confirmed by immunofluorescence microscopy and its enzymatic ability to form ${\alpha}$-cyclodextrin from starch. The maximum surface-display of CGTase was obtained by growing recombinant S. cerevisiae at $20^{\circ}C$ and pH 6.0. S. cerevisiae cells displaying CGTase on their surface consumed glucose and maltose, inhibitory byproducts of the CGTase reaction, to enhance the purity of produced cyclodextrins. Accordingly, the experimental results described herein suggest a possibility of using the recombinant S.cerevisiae anchored with bacterial CGTase on the cell surface as a whole-cell biocatalyst for the production of cyclodextrin.

• Journal of Microbiology and Biotechnology
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• 제25권12호
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• pp.2016-2023
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• 2015

Xylanase plays important roles in a broad range of industrial production as a biocatalyst, and its applications commonly require immobilization on supports to enhance its stability. Aluminum hydroxide, a carrier material with high surface area, has the advantages of simple and low-cost preparation and resistance to biodegradation, and can be potentially used as a proper support for xylanase immobilization. In this work, xylanase from Thermomyces lanuginosus was immobilized on two types of aluminum hydroxide particles (gibbsite and amorphous Al(OH)₃) through adsorption, and the properties of the adsorbed enzymes were studied. Both particles had considerable adsorptive capacity and affinity for xylanase. Xylanase retained 75% and 64% of the original catalytic activities after adsorption to gibbsite and amorphous Al(OH)₃. Both the adsorptions improved pH and thermal stability, lowered activation energy, and extended lifespan of the immobilized enzyme, as compared with the free enzyme. Xylanase adsorbed on gibbsite and amorphous Al(OH)₃ retained 71% and 64% of its initial activity, respectively, after being recycled five times. These results indicated that aluminum hydroxides served as good supports for xylanase immobilization. Therefore, the adsorption of xylanase on aluminum hydroxide particles has promising potential for practical production.

• Journal of Microbiology and Biotechnology
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• 제20권2호
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• pp.332-339
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• 2010

The lipase from Mucor racemosus NRRL 3631 was partially purified by fractional precipitation using 60% ammonium sulfate, which resulted in a 8.33-fold purification. The partially purified lipase was then immobilized using different immobilization techniques: physical adsorption, ionic binding, and entrapment. Entrapment in a 4% agar proved to be the most suitable technique (82% yield), as the immobilized lipase was more stable at acidic and alkaline pHs than the free enzyme, plus 100% of the original activity was retained owing to the thermal stability of the immobilized enzyme after heat treatment for 60 min at $45^{\circ}C$. The calculated half-lives (472.5, 433.12, and 268.5 min at 50, 55, and $60^{\circ}C$, respectively) and the activation energy (9.85 kcal/mol) for the immobilized enzyme were higher than those for the free enzyme. Under the selected conditions, the immobilized enzyme had a higher $K_m$ (11.11 mM) and lower $V_{max}$ (105.26 U/mg protein) when compared with the free enzyme (8.33 mM and 125.0 U/mg protein, respectively). The operational stability of the biocatalyst was tested for both the hydrolysis of triglycerides and esterification of fatty acids with glycerol. After 4 cycles, the immobilized lipase retained approximately 50% and 80% of its original activity in the hydrolysis and esterification reactions, respectively.

• Journal of Microbiology and Biotechnology
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• 제20권2호
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• pp.325-331
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• 2010

Rhodococcus erythropolis amidase was expressed in Escherichia coli cells. The crude amidase in the cell-free extract was immobilized using the cross-linked enzyme aggregate (CLEA) method. The crude amidase was mixed with bovine serum albumin and then precipitated with ammonium sulfate. The resultant precipitant was subsequently cross-linked with glutaraldehyde. Scanning electron microscopy revealed that this co-CLEA had a ball-like shape with a diameter of approximately $1\;{\mu}m$. This co-CLEA evidenced hydrolytic activity toward a variety of amide substrates. The amidase co-CLEA evidenced an optimum temperature of $60^{\circ}C$ and an optimum pH of 8.0, results that were similar to those of the soluble amidase. The reaction stability of the co-CLEA was increased. That is, it was stable up to $50^{\circ}C$ and in a pH range of 5.0-12.0. Additionally, the co-CLEA could be recovered by centrifugation, and retained 96% activity after 3 repeated cycles. This amidase co-CLEA may prove useful as a substitute for soluble amidase as a biocatalyst in the pharmaceutical and chemical industries.

• Journal of Microbiology and Biotechnology
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• 제24권10호
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• pp.1382-1388
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• 2014

Proteus vulgaris K80 lipase was expressed in Escherichia coli BL21 (DE3) cells and immobilized on amine-terminated magnetic microparticles (Mag-MPs). The immobilization yield and activity retention were 84.15% and 7.87%, respectively. A homology model of lipase K80 was constructed using P. mirabilis lipase as the template. Many lysine residues were located on the protein surface, remote from active sites. The biochemical characteristics of immobilized lipase K80 were compared with the soluble free form of lipase K80. The optimum temperature of K80-Mag-MPs was $60^{\circ}C$, which was $20^{\circ}C$ higher than that of the soluble form. K80-Mag-MPs also tended to be more stable than the soluble form at elevated temperatures and a broad range of pH. K80-Mag-MP maintained its stable form at up to $40^{\circ}C$ and in a pH range of 5.0-10.0, whereas soluble K80 maintained its activity up to $35^{\circ}C$ and pH 6.0-10.0. K80-Mag-MPs had broader substrate specificity compared with that of soluble K80. K80-Mag-MPs showed about 80% residual relative activity after five recovery trials. These results indicate the potential benefit of K80-Mag-MPs as a biocatalyst in various industries.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.540-543
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• 2000

본연구에서는 Rhodococcus rhodochrous IGTS8 균주를 사용하여 고농도의 유상에서 탈황효율을 높이기 위한 오일 함유비, pH, 영양물질의 영향을 조사하였다. 오일 함유비에 의한 영향은 유상이 30%이하일 경우 그리 크지 않았으며, pH 조절에 의해 50%, 영양물질의 강화에 의해 32%의 탈황효율이 증가했다. 강화배지에서 pH를 조절하며 배양한 결과, 기존의 배양에 비해 136% 탈황효율이 증가했다.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.191-193
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• 2000

B. macerans 유래의 CGTase를 yeast surface display기술을 이용하여 S. cerevisiae의 표면에 발현된 것을 halo-test와 immunofluorescence microscopy와 flow cytometry를 통하여 확인하였다. 재조합 효모는 효소의 cyclization작용을 저해하고 CD의 분해작용을 촉진하는 glucose와 maltose를 제거하는 발효공정과 표면 발현된 CGTase의 cyclization 공정을 동시에 수행할 수 있어 CD의 생산, 분리공정을 효율적으로 개선하였다.

• Korean Chemical Engineering Research
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• 제54권6호
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• pp.817-821
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• 2016

Levodopa or L-3,4-dihydroxyphenylalanine (L-DOPA) is the direct precursor of the neurotransmitter dopamine. L-DOPA is a well-known neuroprotective agent for the treatment of Parkinson's disease symptoms. L-DOPA was synthesized using the enzyme, tyrosinase, as a biocatalyst for the conversion of L-tyrosine to L-DOPA and an electrochemical method for reducing L-DOPAquinone, the product resulting from enzymatic synthesis, to L-DOPA. In this study, three electrode systems were used: A glassy carbon electrode (GCE) as working electrode, a platinum, and a Ag/AgCl electrode as auxiliary and reference electrodes, respectively. GCE has been modified using electropolymerization of pyrrole to facilitate the electron transfer process and immobilize tyrosinase. Optimum conditions for the electropolymerization modified electrode were a temperature of $30^{\circ}C$ and a pH of 7 producing L-DOPA concentration 0.315 mM. After 40 days, the relative activity of an enzyme for electropolymerization remained 38.6%, respectively.

• KSBB Journal
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• 제10권2호
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• pp.159-165
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• 1995

현탁상 효소반응기 (suspended bed enzyme reac­t tor)에서 alginate-enclosed microspheres를 이용 하여 에틸 프룩토시드의 연속생산과 그에 따른 반융 기 모사로부터 조업 조건의 최적화를 검토하고 메탈 프룩토시드의 연속생산 공정에 관한 체계적인 자료 를 제시하여 효소법에 의한 메틸 프룩토시드 연속생 산의 상엽적 실용성을 제고하고자 하였다. 그 결과, 자당농도 $20.291mol/\ell$, 반응온도 $25^{\circ}C$.에서 최척 에 탄올 함량은 30% (v/v), 최적 pH는 4.8, 최적 효소 활성도는 2U/ml로 나타났으며, 연속생산으로 얻을 수 있는 메틸 프룩토시드의 최대 농도는 $28.4g/\ell$이며, 에틸 프룩토시드의 생산성과 놓도를 함께 고려 한 최척 공간 속도는 $0.1hr^{-1}$부근으로 판단되었고, 그때의 생산성은 $2g/\ell-hr$이며, 그 수율은 47.1 %이 였다.

• 대한화학회지
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• 제36권5호
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• pp.753-759
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• 1992

흰 무궁화꽃의 씨방을 이용하여 Tissue electrode를 제조하고, L-arginine을 정량하기 위한 최적조건을 조사한 결과 0.1M phosphate 완충용액에서 pH 8.0, 온도 $25^{\circ}C$, 조직두께 $20 {\mu}m$이었을 때이며, 이에 정량 가능한 직선범위는 $5.0 {\times} 10^{-3}M $∼ $1.0 {\times} 10^{-1}M$이었으며, 감응도는 54.0 mV/decade로 나타났고, 감응시간은 7∼8분이 소요되었다. $K_m$값은 $6.4 {\times} 10^{-6}(M)$이다.