Developing Bacillus subtilis as a versatile bioproduct platform for agricultural and pharmaceutical applications

Bioproducts made by microbial cell factories play important roles in human life ranging from food, feed, cosmetic and pharmaceutical manufacturing. Microbial hosts are required to be non-pathogenic overproducers and should be growing at low costs. Bacillus subtilis is one of the bacteria, which can fulfill these requirements. It is generally regarded as safe, easy to modify genetically and able to grow fast in simple medium. It can secrete high levels of proteins into the culture medium, and has also been reported to be a high isoprene producer. The work described in the thesis of Yafeng Song focuses on two aspects. Firstly, she engineered B. subtilis to secrete high level of β-mannanase, a major mannan-degrading enzyme that is extensively used in agriculture as feed additive. This was achieved by increasing β-mannanase synthesis levels and enhancing protein secretion capacity. Secondly, she engineered B. subtilis to produce high levels of squalene and amorphadiene. Squalene is a critical precursor of many bioactive triterpenoids, and itself can also be applied as a pharmaceutical. Biosynthesis of squalene and its production improvement in B. subtilis was realized mainly by screening suitable squalene synthases, and increasing precursor supply. Amorphadiene is the key intermediate of the antimalarial drug artemisinin. Its production was elevated by systematically engineering the precursor supply pathway, competitive pathways, terpene synthase and a central metabolic pathway of B. subtilis. Overall, studies in this thesis demonstrated the potential of B. subtilis as a versatile bioproduct platform and provide universal strategies to engineer B. subtilis for industrial applications.