• 분석과학
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• 제12권6호
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• pp.565-570
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• 1999

Bacillus stearothermopilus의 염색체 DNA로부터 polymerase chain reaction 법을 이용하여 ADH의 구조유전자를 증폭시킨 후, 발현 벡터 pGEX-KC에 삽입시켜 glutathion S-tansferase와 융합 단백질로 대장균에서 대랑 발현시켰다. 재조합 ADH는 $37^{\circ}C$에서 1 mM의 isopropyl-${\beta}$-D-thiogalactopyranoside로 단백질의 발현을 유도시켰으며, 발현된 단백질은 glutathione affinity 컬럼을 이용하여 정제하였다. 재조합 ADH는 에탄올에 높은 기질특이성을 나타내었으며 최적 pH와 온도는 각각 pH 9.0과 $70^{\circ}C$이었다. 또한 이 재조합 ADH는 본래의 효소보다 열에 안정하였다. 이 효소는 알코을 측정을 위한 효소학적 방법과 알코올의 공업적 생산에 이용될 수 있다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제10권4호
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• pp.329-333
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• 2005

This study was designed to investigate the stability of a lipase fused with a cellulose­binding domain (CBD) to cellulase. The fusion protein was derived from a gene cluster of a CBD fragment of a cellulase gene in Trichoderma hazianum and a lipase gene in Bacillus stearother­mophilus L1. Due to the CBD, this lipase can be immobilized to a cellulose material. Factors affecting the lipase stability were divided into the reaction-independent factors (RIF), and the re­action-dependent factors (RDF). RIF includes the reaction conditions such as pH and tempera­ture, whereas substrate limitation and product inhibition are examples of RDF. As pH 10 and $50^{\circ}C$ were found to be optimum reaction conditions for oil hydrolysis by this lipase, the stability of the free and the immobilized lipase was studied under these conditions. Avicel (microcrystal­line cellulose) was used as a support for lipase immobilization. The effects of both RIF and RDF on the enzyme activity were less for the immobilized lipase than for the free lipase. Due to the irreversible binding of CBD to Avicel and the high stability of the immobilized lipase, the enzyme activity after five times of use was over $70\%$ of the initial activity.