• KSBB Journal
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• v.18 no.4
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• pp.306-311
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• 2003

Solid-phase refolding of immobilized proteins can be an effective way to reuse an immobilized enzyme column. Oriented immobilization methods are known to provide higher activity of the immobilized enzymes. In this study, using recombinant EK (enterokinase) as a model enzyme and a fusion protein, that consisted of recombinant human growth hormone and six His tag that was linked by the peptide of EK-specific recognition sequence, as a model substrate, we evaluated two oriented immobilization methods, i. e., reductive alkylation of N-terminus ${\alpha}$-amine and affinity interaction between poly-histidine tag and Ni-NTA (nickel-nitrilotriacetic acid). The immobilization yield, activity and cleavage of the immobilized enzymes, and the yield of solid-phase refolding were compared. The Ni affinity immobilization and the covalent immobilization yields were about 100％ and 65％, respectively. But the specific activities were the same, about 50％ of that of the soluble enzyme. The cleavage rate by the covalently immobilized EK was higher than the soluble enzyme and the side reaction of cryptic cleavage was significantly decreased. Covalently immobilized EK showed almost 100％ refolding yield but the affinity immobilized EK showed only 70％ yield, which suggested the covalent conjugation provided more rigid ‘reference structure’ for the solid-phase refolding. The monomeric hGH could be easily obtained by capturing the cleaved poly Histidine tag by the Ni affinity column.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.1043-1046
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• 2004

The stability and electrochemical properties of a self-assembled layer of spinach ferredoxin on a quartz substrate and on a highly oriented pyrolytic graphite electrode were investigated. To fabricate the ferredoxin self-assembly layer, dimyristoylphosphatidylcholine was first deposited onto a substrate for ferredoxin immobilization. Surface analysis of the ferredoxin layer was carried out by atomic force microscopy to verify the ferredoxin immobilization. To verify ferredoxin immobilization on the lipid layer and to confirm the maintenance of redox activity, absorption spectrum measurement was carried out. Finally, cyclic-voltammetry measurements were performed on the ferredoxin layers and the redox potentials were obtained. The redox potential of immobilized ferredoxin had a formal potential value of -540 mV. It is suggested that the redox-potential measurement of self-assembled ferredoxin molecules could be used to construct a biosensor and biodevice.

• Korean Chemical Engineering Research
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• v.44 no.1
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• pp.65-72
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• 2006

Recent technical advances in the biorecognition engineering and the microparticle fabrication may enable us to develop the single step purification using magnetic particle, because of its simplicity, efficacy, ease of automation, and process economics. In this study, we used commercial magnetic particles from Seradyn, Inc. (Indianapolis, USA). It was ca. 2.8 micron in diameter, consisted of polystyrene core and magnetite coating, and its surface had carboxyl groups. The model, capture protein was IgG and anti-IgG was used as the ligand molecule. We studied the different surfaces ('nude', ester-activated, and anti-IgG coated) for their biorecognition of IgG. At a high pH condition, we could reduce non-specific binding. Also anti-IgG immobilized magnetic particle could capture IgG more selectively. We attempted 'oriented immobilization' of anti-IgG, in which the polysaccharides moiety near the C-terminus was selectively oxidized and linked to the hydrazine-coated MP, to improve the efficacy of biorecognitive binding. Using this method, the IgG capturing ability was improved by ca. 2 fold. From the binary mixture of the IgG-insulin, IgG could be more selectively captured. In summary, the oriented immobilization of oxidized anti-IgG proved to be as effective as the streptavidin-biotin system and yet simpler and cost-effective. This immobilization method can find its applications in protein biochips and biotargeting.

• Journal of Korea Technology Innovation Society
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• v.16 no.4
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• pp.1134-1162
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• 2013

The purpose of this paper is to provide an innovation strategy for a leading industrial company. We analyze the process of a shift in industrial leadership using the shipbuilding industry as a case study. This paper explains that the three core factors of decline are immobilization, lack of innovation for major products, and organizational inertia. This study presents that the leader of a market is not decided by strong competitiveness, but by quick reaction to major customers' needs, and that out-of-date strategies do not contribute to maintaining leadership. As a result of this study, it shows that the leading Korean industrial companies shall establish a new innovation-oriented and customer-oriented management system that is flexible enough to anticipate and respond to the changes in new competitive environments. These findings contribute to the formulation of new strategies for other manufacturing industries in Korea.