Creation of an orthogonal and universal auto-inducible gene expression platform by reprogramming a two-component signal circuit for efficient production of industrial enzymes.

Abstract

Bacterial gene expression systems play a crucial role in producing valuable biological macromolecules, such as recombinant proteins and polysaccharides. However, traditional inducible gene systems have limitations that need costly chemical inducers that can harm the host. To address these challenges, a novel peptide-activated auto-inducible gene expression system was developed in Bacillus subtilis, leveraging Accessory gene regulatory system (Agr), a two-component signal system, from Staphylococcus aureus to trigger gene expression in response to an auto-inducible peptide (AIP). This system mimics a cell density-dependent regulatory mechanism, allowing for the intuitive activation of gene expression as accumulation of AIP. By precisely tuning the level of AIP, the auto-induction time was successfully delayed, however, at the expense of slightly reducing the strength of effector promoter P3, thus decreasing level of output expression. Furthermore, modulation of the stoichiometry of sensor protein AgrC allowed for fine-tuning of the auto-induction time, temporal dynamics, and expression levels. The robustness of the system was improved by strengthening P3 while maintaining the delayed auto-induction time. The versatility and efficacy of the system was demonstrated by the efficient production of various industrial enzymes. This study paves the way for the application of bacterial two-component signal systems to design synthetic gene circuits.