Increased production of riboflavin by metabolic engineering of the purine pathway in Bacillus subtilis

Purine nucleotides are precursors for riboflavin production, and one of its intermediates, GTP, is the immediate precursor for riboflavin biosynthesis. Therefore, activation of the purine pathway would improve the supply of the precursors, which may in turn lead to an increase of riboflavin production. Here we reported the characterization and manipulation of the purine pathway in a riboflavin overproducer Bacillus subtilis PK. Due to the tight regulation of the purine pathway, it was not sufficient to obtain an enhanced production of purine nucleotides or riboflavin by simple overexpression of the purF gene, which is the key gene of the purine pathway. However, co-overexpression of purF, purM, purN, purH, purD genes gave an engineered strain B. subtilis PK-P with much better performance. Metabolites analysis revealed that the precursors, purine nucleotides, were significantly improved in B. subtilis PK-P. Moreover, as much as a 25% increase in the riboflavin yield and a 31% increase in riboflavin titer were obtained in the recombinant B. subtilis PK-P. It was assumed that the co-overexpression of the genes caused higher supply of the purine nucleotides in B. subtilis PK-P, which in turn facilitated riboflavin production.