Abstract

An additional amylase, besides the typical a-amylase, was detected for the first time in the cytoplasm of B. subtilis SUH4-2, an isolate from Korean soil. The corresponding gene (bbmA) encoded a maltogenic amylase (MAase) and its sequence was almost identical to the yvdF gene of B. subtilis 168, whose function was unknown. Southern blot analysis using bbmA as the probe indicated that this gene was ubiquitous among various B. subtilis strains. In an effort to understand the physiological function of the bbmA gene in B. subtilis, the expression pattern of the gene was monitored by measuring the [beta]-galactosidase activity produced from the bbmA promoter fused to the amino terminus of the lacZ structural gene, which was then integrated into the amyE locus on the B. subtilis 168 chromosome. The promoter was induced during the mid-log phase and fully expressed at the early stationary phase in defined media containing [beta]-cyclodextrin ([beta]-CD), maltose, or starch. On the other hand, it was kept repressed in the presence of glucose, fructose, sucrose, or glycerol, suggesting that catabolite repression might be involved in the expression of the gene. Production of the [beta]-CD hydrolyzing activity was impaired by the spo0A mutation in B. subtilis 168, indicating the involvement of an additional regulatory system exerting control on the promoter. Inactivation of yvdF resulted in a significant decrease of the [beta]-CD hydrolyzing activity, if not all. This result implied the presence of an additional enzyme(s) that is capable of hydrolyzing [beta]-CD in B. subtilis 168. Based on the results, MAase encoded by bbmA is likely to be involved in maltose and [beta]-CD utilization when other sugars, which are readily usable as an energy source, are not available during the stationary phase.

• Keywords: Maltogenic amylase; B. subtilis; yvdF; cyclodextrin utilization; catabolite repression; spo0A