Cloning of the Entire Gene Encoding the 140-kDa $\alpha$-Amylase of Lactobacillus amylovorus and Expression in Escherichia coli and Lactococcus lactis

A 4.6-kb HindIII fragment encompassing the complete 140-kDa -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 -amylase activity to E. coli and Lactococcus lactis hosts when introduced by electroporation. SDS-PAGE and zymography confirmed the production of 140-kDa -amylase and its proteolytic degradation products with enzyme activity in transformants. Total -amylase activity of E. coli 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.