Engineering of a Xylose Metabolic Pathway in Corynebacterium glutamicum

The aerobic microorganismCorynebacterium glutamicum was metabolically engineered to broaden its substrate utilization range to include the pentose sugar xylose, which is commonly found in agricultural residues and other lignocellulosic biomass. We demonstrated the functionality of the corynebacterialxylB gene encoding xylulokinase and constructed two recombinantC. glutamicum strains capable of utilizing xylose by cloning theEscherichia coli genexylA encoding xylose isomerase, either alone (strain CRX1) or in combination with theE. coli genexylB (strain CRX2). These genes were provided on a high-copy-number plasmid and were under the control of the constitutive promotertrc derived from plasmid pTrc99A. Both recombinant strains were able to grow in mineral medium containing xylose as the sole carbon source, but strain CRX2 grew faster on xylose than strain CRX1. We previously reported the use of oxygen deprivation conditions to arrest cell replication inC. glutamicum and divert carbon source utilization towards product production rather than towards vegetative functions (M. Inui, S. Murakami, S. Okino, H. Kawaguchi, A. A. Vertès, and H. Yukawa, J. Mol. Microbiol. Biotechnol.7 :182-196, 2004). Under these conditions, strain CRX2 efficiently consumed xylose and produced predominantly lactic and succinic acids without growth. Moreover, in mineral medium containing a sugar mixture of 5% glucose and 2.5% xylose, oxygen-deprived strain CRX2 cells simultaneously consumed both sugars, demonstrating the absence of diauxic phenomena relative to the newxylA-xylB construct, albeit glucose-mediated regulation still exerted a measurable influence on xylose consumption kinetics.