Transcriptomic analysis of engineered and evolved Klebsiella oxytoca with efficient glucose/xylose co-utilization and...

Klebsiella oxytoca strain KC004-TF160 (ΔadhEΔpta-ackAΔldhAΔbudABΔpflB) was previously engineered as succinate-producing biocatalysts. To further improve glucose/xylose co-utilization, KC004-XY70 was developed through adaptive evolution of KC004-TF160, resulting in enhanced xylose utilization and increased co-consumption of glucose and xylose up to 44.4 %. Based on transcriptomes, significant regulatory changes included downregulation of xylFGH and xylR, reducing ATP expenditure during ABC transporter-mediated xylose uptake, and upregulation of gudP and garP (galactarate/d-glucarate transporters), enabling xylose import via major facilitator superfamily (MFS). Although expression of genes in pentose phosphate pathway and glycolysis was reduced, xylose consumption and growth remained unaffected, suggesting that lower enzymatic activities sufficed for efficient metabolism. KC004-XY70 achieved 84.2, 61.9, and 71.3 g/L succinate from glucose, xylose, and their mixtures, representing a 30.5-36.1 % increase over KC004-TF160. Moreover, succinate yields up to 0.83 g/g sugars consumed were obtained from NaOH-pretreated Napier grass, underscoring the potential by KC004-XY70 for industrial-scale succinate production from other lignocellulosic feedstocks.