Riboflavin overproduction from diverse feedstocks with engineered<i>Corynebacterium glutamicum</i>.

Abstract

Riboflavin overproduction by<i>Corynebacterium glutamicum</i>was achieved by screening synthetic operons, enabling fine-tuned expression of the riboflavin biosynthetic genes<i>ribGCAH.</i>The synthetic operons were designed by means of predicted translational initiation rates of each open reading frame, with the best-performing selection enabling riboflavin overproduction without negatively affecting cell growth. Overexpression of the fructose-1,6-bisphosphatase (<i>fbp</i>) and 5-phosphoribosyl 1-pyrophosphate aminotransferase (<i>purF</i>) encoding genes was then done to redirect the metabolic flux towards the riboflavin precursors. The resulting strain produced 8.3 g l^-1of riboflavin in glucose-based fed-batch fermentations, which is the highest reported riboflavin titer with<i>C. glutamicum</i>. Further genetic engineering enabled both xylose and mannitol utilization by<i>C. glutamicum</i>, and we demonstrated riboflavin overproduction with the xylose-rich feedstocks rice husk hydrolysate and spent sulfite liquor, and the mannitol-rich feedstock brown seaweed hydrolysate. Remarkably, rice husk hydrolysate provided 30% higher riboflavin yields compared to glucose in the bioreactors.