대장균에서의 Chaperone 동시 발현을 통한 Candida antarctica Lipase B 발현 시스템 구축

Abstract

Recently, Candida antarctica lipase B (CalB) draws attention from industries for various applications for food, detergent, fine chemical, and biodiesel, because of its characteristics as an efficient biocatalyst. Since many industrial processes carry out in organic solvent and at high temperature, CalB, which is stable under harsh condition, is in demand from many industries. In order to reform CalB promptly, the expression system which has advantages of ease to use and low cost for gene libraries screening was developed using E. coli. The E. coli strains, Rosettagami with competence for enhanced disulfide bond formation, Novablue, and DH5α, were exploited in this study. To obtain the soluble CalB, the pCold I vector expressing the cloned gene at 15℃ and the chaperone plasmids containing groES/groEL, groES/groEL/tig, tig, dnaK/dnaJ/grpE, and dnaK/dnaJ/grpE/groES/groEL were used for coexpression of CalB and chaperones. The colonies expressing functional lipase were selected by employing the halo plate containing 1% tributyrin, and the CalB expression was confirmed by SDS-PAGE. E. coli Rosettagami and DH5α harbouring groES/groEL chaperones were able to express soluble CalB effectively. From a facilitative point of view, E. coli DH5α is more suitable for further mutation study.