KMID : 0545119970070050293
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Journal of Microbiology and Biotechnology 1997 Volume.7 No. 5 p.293 ~ p.298
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Cloning of the Entire Gene Encoding the 140-kDa ¥á-Amylase of Lactobacillus amylovorus and Expression in Escherichia coli and Lactococcus lactis
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Jeong, Jong-Jin
Kim, Tea-Youn/Kim, Jeong Hwan
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Abstract
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A 4.6-kb Hind¥² fragment encompassing the complete 140-kDa ¥á-amylase gene of Lactobacillus amylovorus B 4540 was cloned into pBR322 by the shot gun method. Southern blotting and restriction mapping for the insert were performed. The recombinant 9.0-kb plasmid, pFML1, conferred ¥á-amylase activity to E. coli and Lactococcus lactis hosts when introduced by electroporation. SDS-PAGE and zymography confirmed the production of 140-kDa ¥á-amylase and its proteolytic degradation products with enzyme activity in transformants. Total ¥á-amylase activity of E. coli DH5¥á cells harboring pFML1 was 1.8 units and most activity was detected from cell pellets. Total enzyme activity of L. lactis subsp. lactis MG1363 transformant was five to ten-fold lower than that of E. coli cell but more than half of the activity was detected in the culture supernatant.
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