Fig. 1.

Schematic representation of strategies in developing the MATE system in B. subtilis. Different expression elements, including the promoter core region, cis-element, translational riboswitch, and chassis cell, were combinatorially optimized to generate the inducible MATE-ON and MATE-OFF systems with robust, stringent, and homogeneous characteristics. The mal operon in B. subtilis, which is responsible for the uptake and hydrolysis of the inducer maltose, was engineered to improve the sensitivity, long-term induction, and homogeneity of our system. The strength of the PmalA promoter was elevated by mutagenesis and high-throughput screening, and the stringency was improved by modifying the cre-element and introducing a translational riboswitch device. Finally, the universality and compatibility of our system were demonstrated by different applications in protein expression and synthetic biology.