[Efficient biosynthesis of spermidine via whole-cell catalysis with a multi-enzyme cascade system]

Spermidine is a natural polyamine with autophagy-inducing properties, capable of extending lifespan, inhibiting tumors, protecting cardiovascular health, regulating neural functions, and exerting anti-inflammatory effects. It holds significant applications in agriculture, industry, and healthcare. This study focuses on the microbial synthesis of spermidine by tracing the biosynthetic pathways of its precursors. Through the overexpression of key enzymes involved in precursor synthesis, spermidine was successfully synthesized in vitro with amino acids as substrates under multi-enzyme cascade conditions, which significantly reduced the production costs. The phylogenetic analysis identified an efficient S-adenosylmethionine decarboxylase from Bacillus subtilis, and its activity was enhanced by 2.68 folds through mutagenesis. Subsequently, the superiority of whole-cell catalysis for spermidine synthesis was confirmed, and the continuous and efficient synthesis of spermidine was achieved by optimizing gene tandem sequences and introducing an ATP regeneration system. Further, the reaction conditions, including cell density, substrate concentration, reaction temperature, pH, and metal ions, were optimized. Under optimal conditions, the recombinant Escherichia coli EC12 achieved a maximum spermidine yield of 17.83 g/L in a 5 L bioreactor after 20 hours of transformation. The recombinant E. coli constructed in this study demonstrates potential for industrial-scale spermidine production, laying a theoretical and practical foundation for the green and efficient synthesis of spermidine.