Flexibility with Truncation of the Amino- and Carboxyl-termini of Bacillus subtilis X-23 .ALPHA.-Amylase.

Abstract

We analyzed the amino- and carboxyl-terminal amino acid sequences of five .α-amylases, Ba-L, Ba-S, pUXAI Amy, pUXA0 Amy, and pBLXA1 Amy, which originate from an α-amylase gene of Bacillus subtilis X-23. Ba-L and Ba-S were produced by wild-type strain B. subtilis X-23, while pUXAI Amy, pUXA0 Amy and pBLXA1 Amy were produced by Escherichia coli harboring dif ferent kinds of recombinant plasmids coding the α-amylase gene. Amino-terminal amino acid se quence analyses indicated that the signal peptides of Ba-L and Ba-S were the first 45 amino acid residues and those of pUXA1 Amy and pUXA0 Amy were the first 31 amino acid residues, while that of pBLXA1 Amy was the first 37 amino acid residues. Carboxyl-terminal sequence analyses showed that Ba-L and pUXA1 Amy contained the full length of the carboxyl-terminal-region polypeptide of the enzyme while Ba-S lacked 186 amino acid residues, and that pUXA0 Amy and pBLXA1 Amy lacked 88 and 189 amino acid residues, respectively, but had an extra 8 and 64 amino acid residues, respectively, originating from continuous translation of the vector region. All five enzymes showed the same characteristics with regard to molar activity, optimum temperature, optimum pH, and transglycosylation activity. These findings suggest that the amino- and carboxyl terminal regions of α-amylase have no effect on enzyme activity, but rather are flexible with regard to the truncation and addition of polypeptides.