Structure and expression of a gene coding for thermostable α‐glucosidase with a broad substrate specificity from Bacillus sp. SAM1606

We cloned an α‐glucosidase gene from thermophilic Bacillus sp. SAM1606 to overexpress it in Escherichia coli transformants. Deletion of the 5′‐noncoding region as well as expression of the α‐glucosidase gene under the control of the icp promotor of the insecticidal crystal protein gene from Bacillus thuringiensis subsp. sotto enhanced the enzyme productivity to 23.5 U/ml, which was 12000‐fold higher than that obtained by the strain SAM1606. The open reading frame corresponding to the α‐glucosidase encoded 587 amino acid residues including a residue coded by the initiation codon TTG, and the molecular mass of the α‐glucosidase from N‐terminal serine was calculated to be 68886Da. Sequence analysis revealed the SAM1606 α‐glucosidase showed extremely high sequence identity (62–65%) to the Bacillus cereus and Bacillus thermoglucosidasius oligo‐1,6‐glucosidases, which were 72% identical to each other. Sequence identity in the suggested active site regions were essentially the same (80–82%) among these three enzymes. However, the substrate specificity of the SAM1606 α‐glucosidase was significantly different from those of the oligo‐1,6‐glucosidases. The thermostability of these three α‐glucosidases could be correlated with the increase in the number of proline residues, whose occurrence was predicted at β turns and coils in the enzymes.