Bacterial amylase genes and their expression.

We cloned genes coding for two thermophilic α-amylases from Bacillus licheniformis and B. stearothermophilus and one β-amylase from B. polymyxa. The nucleotide sequence determination of these genes revealed the characteristics of their sequences such as promoter and signal sequence were clarified. The thermostability of the thermophilic amylases was discussed on the basis of the hydropathy profile of amino acid sequence deduced from the nucleotide sequence. The recombinant DNA technology was shown to be useful for determining the region responsible for the thermostability or optimal pH of the enzymes. Amino acid sequences of liquefying α-amylases secreted by the bacteria of Bacillus species were found to be similar each other. Interestingly, three homologous sequences at the active sites (catalytic and substrate binding sites) were conserved among not only bacterial amylases but also amylases of fungal and animal origins. The thermophilic amylase genes were expressed in heterologous bacteria such as Escherichia coli and B. subtilis. Most efficient formation and secretion of the enzyme (as high as 0.5g/liter) were observed when B. brevis 47, a protein-producing bacterium, was used as a host. β-Amylases with three different molecular weights were produced by both B. polymyxa and heterologous bacteria having the β-amylase gene on plasmids. These amylases have exactly the same amino acid sequence at the amino terminal region and cross-reacted with the antibody raised for the β-amylase with the largest molecular weight.