Abstract
BackgroundSuccinate has been recognized as one of the most important bio-based building block chemicals due to its numerous potential applications. However, efficient methods for the production of succinate from lignocellulosic feedstock were rarely reported. Nevertheless, Corynebacterium glutamicum was engineered to efficiently produce succinate from glucose in our previous study.ResultsIn this work, C. glutamicum was engineered for efficient succinate production from lignocellulosic hydrolysate. First, xylose utilization of C. glutamicum was optimized by heterologous expression of xylA and xylB genes from different sources. Next, xylA and xylB from Xanthomonas campestris were selected among four candidates to accelerate xylose consumption and cell growth. Subsequently, the optimal xylA and xylB were co-expressed in C. glutamicum strain SAZ3 (ΔldhAΔptaΔpqoΔcatPsod-ppcPsod-pyc) along with genes encoding pyruvate carboxylase, citrate synthase, and a succinate exporter to achieve succinate production from xylose in a two-stage fermentation process. Xylose utilization and succinate production were further improved by overexpressing the endogenous tkt and tal genes and introducing araE from Bacillus subtilis. The final strain C. glutamicum CGS5 showed an excellent ability to produce succinate in two-stage fermentations by co-utilizing a glucose–xylose mixture under anaerobic conditions. A succinate titer of 98.6 g L−1 was produced from corn stalk hydrolysate with a yield of 0.87 g/g total substrates and a productivity of 4.29 g L−1 h−1 during the anaerobic stage.ConclusionThis work introduces an efficient process for the bioconversion of biomass into succinate using a thoroughly engineered strain of C. glutamicum. To the best of our knowledge, this is the highest titer of succinate produced from non-food lignocellulosic feedstock, which highlights that the biosafety level 1 microorganism C. glutamicum is a promising platform for the envisioned lignocellulosic biorefinery.
Highlights
Succinate has been recognized as one of the most important bio-based building block chemicals due to its numerous potential applications
Selection of optimal xylose isomerases and xylulokinases for heterologous expression Heterologous expression of the XI pathway from E. coli endowed C. glutamicum with the ability to grow on xylose [19, 23]
The xylose isomerase (xylA) and xylB genes from E. coli MG1655, P. polymyxa SC2, S. coelicolor, and X. campestris 8004 were cloned into the IPTGinducible expression vector pXMJ19
Summary
Succinate has been recognized as one of the most important bio-based building block chemicals due to its numerous potential applications. There seem to be few reports on the efficient production of succinate from non-food lignocellulosic feedstocks, such as corn stalk, sugarcane bagasse, pine-, oak-, or spruce wood. The strain E. coli SD121 was able to produce 57.8 g L−1 succinate from corn stalk hydrolysate with a yield of 0.87 g/g total sugars [8]. In most reports on succinate production from lignocellulosic biomass, the yields were quite good, but the titers were generally limited by the sugar concentration obtained from the hydrolysis of lignocellulosic biomass, making the process uneconomical for further separation and purification. Corn stalk hydrolysates containing high concentrations of glucose and xylose obtained by means of enzymatic hydrolysis were chosen as substrate for succinate production
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