Construction of a High-Expression System in Bacillus through Transcriptomic Profiling and Promoter Engineering

Bacillus subtilis is an ideal host for secretion and expression of foreign proteins. The promoter is one of the most important elements to facilitate the high-level production of recombinant protein. To expand the repertoire of strong promoters for biotechnological applications in Bacillus species, 14 highly transcribed genes based on transcriptome profiling of B. pumilus BA06 were selected and evaluated for their promoter strength in B. subtilis. Consequently, a strong promoter P₂₀₆₉ was obtained, which could drive the genes encoding alkaline protease (aprE) and green fluorescent protein (GFP) to express more efficiency by an increase of 3.65-fold and 18.40-fold in comparison with the control promoter (P_aprE), respectively. Further, promoter engineering was applied to P₂₀₆₉, leading to a mutation promoter (P_2069M) that could increase GFP expression by 3.67-fold over the wild-type promoter (P₂₀₆₉). Moreover, the IPTG-inducible expression systems were constructed using the lac operon based on the strong promoters of P₂₀₆₉ and P_2069M, which could work well both in B. subtilis and B. pumilus. In this study, highly efficient expression system for Bacillus was constructed based on transcriptome data and promoter engineering, which provide not only a new option for recombinant expression in B. subtilis, but also novel genetic tool for B. pumilus.