New xylose transporters support the simultaneous consumption of glucose and xylose in .

Glucose and xylose are two major components of lignocellulose. Simultaneous consumption of glucose and xylose is critical for engineered microorganisms to produce fuels and chemicals from lignocellulosic biomass. Although many production limitations have been resolved, glucose-induced inhibition of xylose transport remains a challenge. In this study, a cell growth-based screening strategy was designed to identify xylose transporters uninhibited by glucose. The glucose pathway was genetically blocked in so that glucose functions only as an inhibitor and cells need xylose as the carbon source for survival. Through adaptive evolution, omics analysis and reverse metabolic engineering, a new phosphoenolpyruvate: carbohydrate phosphotransferase system (PTS) galactitol transporter (GalABC, encoded by , , and genes) that is not inhibited by glucose was identified. Inactivation of adenylate cyclase led to increased expression of the gene, and a point mutation in gene (N13S) further enhanced xylose transport. During the second round of gene mining, AraE and a new ABC transporter (AraFGH) of xylose were identified. A point mutation in the transcription regulator (L156I) caused increased expression of and genes without arabinose induction, and a point mutation in (D223Y) further enhanced xylose transport. These newly identified xylose transporters can support the simultaneous consumption of glucose and xylose and have potential use in producing chemicals from lignocellulose.