CLONING, SEQUENCING, AND EXPRESSION OF A BETA-AMYLASE GENE FROM BACILLUS-CEREUS VAR MYCOIDES AND CHARACTERIZATION OF ITS PRODUCTS

The cloned gene was composed of 1638 bp for coding plus promoter like and SD-like sequences ahead of it. The deduced amino acid sequence had high similarity with known beta-amylases. The N-terminal sequence of the cloned beta-amylase seemed to be a signal peptide. The gene was in troduced into Bacillus subtilis 1A289 using pHY300PLK as a vector and the expressed protein was recovered from the culture media. The enzyme fraction produced was divided into two components upon the DEAE colum n chromatography. The amino acid sequence of one fraction (FrI) was th e same as the mature enzyme, and the other (FrII) lacked the N-termina l amino acid residue (Ala) of the mature enzyme. The kinetic parameter s of the hydrolysis catalyzed by the enzyme component FrI were measure d, and the subsite affinities of the enzyme were evaluated. In conclus ion, it was shown that the recombinant enzyme was the same as the matu re enzyme functionally and proteochemically.