Convergent Rapid Evolution of Fructose Operon Reveals Its Important Role in Sugar Uptake and Its Effect on Lysine Production

The fructose operon is a conserved structure in many bacteria, which is involved in fructose assimilation. We have identified an evolutionary incentive to keep this operon intact in C. glutamicum by discovering that mutations in two of its members not only affect fructose metabolism, but also the metabolism of sucrose and glucose. The implicated genes are ptsF, encoding the fructose specific component of the sugar phosphotransferase system (PTS) and pfkB, encoding a fructose-1-phosphate kinase.

Inactivation of either of these genes affects growth in minimal medium on all three sugars negatively, but suppressor mutants, where both genes are inactive, appear with high frequency, leading to restored or improved growth on these sugars mentioned above. We adapt a genome scale model for C. glutamicum by overlaying transcriptomic data and are able to predict that the hampered growth of the ptsF knock-out strain can be attributed to perturbed expression of sugar PTS genes, which are found to be drastically down-regulated. We are able to validate this prediction that the hampered growth of KO-ptsF and KO-pfkB mutants are caused by different mechanisms.

Furthermore, when the pfkB and ptsF mutations are introduced in a lysine-producing derivative, the lysine yield is found to be enhanced by almost 200% compared to the control strain on fructose medium and even 100 % higher than the yield observed on glucose. By further over-expressing genes in pentose phosphate pathway and down-regulating the TCA cycle, even higher lysine yields are achieved. The results obtained reveal a novel strategy for producing lysine that takes advantage of the diauxic shift observed when growing C. glutamicum on either glucose/fructose or sucrose.