Programming Low-Molecular-Weight Hyaluronic Acid Biosynthesis in Bacillus amyloliquefaciens Via an Autoinducible Molecular Switch

Dynamic metabolic regulation facilitates real-time sensing of pathway states and precise redirection of metabolic flux, thereby enhancing the biosynthesis of diverse value-added compounds. Here, we designed an autoinducible molecular switch in Bacillus amyloliquefaciens by integrating a Pantoea alhagi -derived PaSspB-ssrA* proteolytic system with the DegU-RapG-PhrG quorum-sensing (QS) module. The OFF module employs the novel PaSspB-ssrA* system for targeted degradation of ssrA*-tagged proteins with 86% efficiency, and the ON module utilizes a DegU-activated P ispA promoter for cell density-dependent gene expression. To improve hyaluronic acid (HA) production and reduce byproduct accumulation, this system was applied by fusing ssrA* tags into the glycolysis-critical genes pfkA and fruA to attenuate glycolytic flux, together with QS-regulated coexpression of PaSspB and SthHL in engineered Δ ldh /Δ wbpA /Δ nagB strains, achieving 13.5 ± 0.19 g/L of low-molecular-weight HA (2.89 × 10 4 Da). The autonomous, inducer-free platform enables cost-effective HA biosynthesis in B. amyloliquefaciens, with broad applicability for synthesizing high-value bioproducts.