Green and sustainable FAME production: Enhanced continuous catalysis based on biofilm surface-displayed enzymes

Fatty acid methyl esters (FAME) are the primary constituents of biodiesel, known for their renewability and environmental benefits. However, during the production of FAME, particularly when homogeneous catalysts are employed, the incomplete separation of the catalyst from the reaction medium poses a significant technical challenge. In this study, we developed an editable, sustainable, and efficient living material for FAME production based on a biofilm system. By overexpressing TasA in Bacillus subtilis we enhanced biofilm formation and created a functional carrier material with lipase activity. The SpyTag/SpyCatcher system immobilizes LipA onto the biofilm surface, improving enzyme stability and reusability. The resulting biofilm displayed a high catalytic capacity for FAME synthesis and could be regenerated to maintain enzyme activity. The living material could catalyze six consecutive batches with a maximum yield of 0.962 nmol mL⁻¹. This approach offers a green and sustainable solution for industrial biocatalysis, aligning with environmental sustainability goals.