Accelerated pentose utilization by Corynebacterium glutamicum for accelerated production of lysine, glutamate, ornithine and putrescine

Abstract

Because of their abundance in hemicellulosic wastes arabinose and xylose are an interesting source of carbon for biotechnological production processes. Previous studies have engineered several Corynebacterium glutamicum strains for the utilization of arabinose and xylose, however, with inefficient xylose utilization capabilities. To improve xylose utilization, different xylose isomerase genes were tested in C. glutamicum. The gene originating from Xanthomonas campestris was shown to have the highest effect, resulting in growth rates of 0.14 h(-1), followed by genes from Bacillus subtilis, Mycobacterium smegmatis and Escherichia coli. To further increase xylose utilization different xylulokinase genes were expressed combined with X. campestris xylose isomerase gene. All combinations further increased growth rates of the recombinant strains up to 0.20 h(-1) and moreover increased biomass yields. The gene combination of X. campestris xylose isomerase and C. glutamicum xylulokinase was the fastest growing on xylose and compared with the previously described strain solely expressing E. coli xylose isomerase gene delivered a doubled growth rate. Productivity of the amino acids glutamate, lysine and ornithine, as well as the diamine putrescine was increased as well as final titres except for lysine where titres remained unchanged. Also productivity in medium containing rice straw hydrolysate as carbon source was increased.

Figure 1

Growth of C. glutamicum strains in CgXII medium containing 100 mM xylose.A. Corynebacterium glutamicum strains WT(pEKEx3-xylA_Xc) (open diamonds), WT(pEKEx3-xylA_Bs) (open triangles), WT(pEKEx3-xylA_Ec) (closed circles) and WT(pEKEx3-xylA_Ms) (closed squares) were analysed.B. Corynebacterium glutamicum strains WT(pEKEx3-xylA_Xc-xylB_Bs) (open diamonds), WT(pEKEx3-xylA_Xc-xylB_Ec) (open triangles), WT(pEKEx3-xylA_Xc) (closed circles) and WT(pEKEx3-xylA_Xc-xylB_Cg) (closed squares) were analysed. Data represents mean values and standard deviations of three independent cultivations.

Figure 2

Product concentrations (A) and volumetric productivities (B) for l-glutamate, l-lysine, l-ornithine and putrescine production in CgXII medium containing 100 mM xylose. Corynebacterium glutamicum strains with pEKEx3-xylA_Ec or pEKEx3-xylA_Xc-xylB_Cg were analysed. l-glutamate was produced with WT (hatched bars), l-lysine with DM1729 (open bars), l-ornithine with ORN1 (closed bars) and putrescine with PUT21 (checked bars). Data represent mean values and experimental imprecision of two independent cultivations.

Figure 3

Product concentrations (A, C) and volumetric productivities (B, D) for l-glutamate (A, B) and l-lysine (C, D) production in CgXII medium containing rice straw hydrolysate. Corynebacterium glutamicum strains with empty vectors, pVWEx1-araBAD and pEKEx3-xylA_Ec or pVWEx1-araBAD and pEKEx3-xylA_Xc-xylB_Cg were analysed. l-glutamate was produced with WT (hatched bars) and l-lysine with DM1729 (open bars). Data represent mean values and experimental imprecision of two independent cultivations.