The low production efficiency of tannase and the insufficient utilization of high-tannin feed resources form the research background and research significance of this study. In this experiment, the tannase sequence TanLpl from Lactiplantibacillus plantarum ATCC14917T (obtained from a microbial culture collection) was selected. These sequences were respectively integrated into the expression systems of Bacillus subtilis 168 (BS168) and Bacillus subtilis WB600 (WB600) through plasmids TanLpl-p43NMK and TanLpl-pHT43. This successfully constructed three tannase-producing strains: TanLpl-p43NMK-Bacillus subtilis 168 (BS168(p43NMK)), TanLpl-pHT43-Bacillus subtilis 168 (BS168(pHT43)), and TanLpl-pHT43-Bacillus subtilis WB600 (WB600(pHT43)). An evaluation of the recombinant strains’ growth characteristics, expression stability, and enzymatic properties revealed that all three strains reached the stationary phase after 18 h of growth, with no significant differences in growth rate compared to the parental strains. At the 10th generation of subculture, the plasmid loss rate of BS168(p43NMK) was significantly higher than that of BS168(pHT43) or WB600(pHT43) (p < 0.05). The optimal temperature for tannase activity in all three recombinant strains was 30 °C, with an optimal pH value of 5.0. Under these conditions, the tannase activities were 68.81 U/mL, 397.36 U/mL, and 461.12 U/mL, respectively. The recombinant strain WB600(pHT43) exhibited superior expression stability and enzyme production capability compared to the other two strains. The research on the heterologous expression of tannase and its application in feed utilization has important theoretical and practical significance: it enriches the technical system for the heterologous expression of functional enzymes in Bacillus subtilis, provides new ideas for the efficient production of industrial enzymes, and promotes the development of bio-manufacturing technology.
Gao et al. (Sat,) studied this question.