High-Titer Glutamic Acid Production from Lignocellulose Using an Engineered Corynebacterium glutamicum with Simultaneous Co-utilization of Xylose and Glucose

Xylose utilization by Corynebacterium glutamicum is an essential but unresolved issue in glutamic acid production from lignocellulose biomass. Coexistence of xylose with inhibitors requires a selective removal of inhibitors while the xylose is still well retained in the pretreated lignocellulose feedstock. Not only is xylose assimilation in C. glutamicum at low efficiency, but also there are unique challenges, which eliminate the generation of glutamic acid from xylose when lignocellulose is used. There include excessive biotin content in lignocellulose blocking intracellular secretion of glutamic acid, complicated organic acid generation pathways decreasing the glutamic acid conversion yield from xylose, and transmembrane resistance of xylose limiting the xylose utilization efficiency. Here, we applied a unique biodetoxification on pretreated wheat straw solids, which resulted in a complete removal of inhibitors and a high conservation of xylose sugar. The major focus of the study is a stepwise metabolic engineering of C. glutamicum to trigger high-titer glutamate production by coordinated assimilation of xylose and glucose from a typical lignocellulosic sugar. First, the secretion channel protein MscCG was modified to initiate glutamic acid secretion in biotin-rich environments from almost zero glutamic acid accumulation. Next, the byproduct generation pathways of lactate, acetate, and succinate were knocked out or attenuated to redirect carbon flux to glutamic acid accumulation. Further overexpression of the pentose transporter gene araE increased the xylose utilization rate and glutamic acid production. The finally obtained C. glutamicum GJ04 produced 39.8 g/L of glutamate from 60.3 g/L of glucose and 38.8 g/L of xylose in synthetic medium and produced 61.7 g/L of glutamate from 116.1 g/L of glucose and 39.6 g/L of xylose using wheat straw feedstock. This is the first example of the practical utilization of lignocellulose-derived xylose and glucose for cellulosic glutamic acid production.