Abstract
BackgroundXylose is the most prevalent sugar available in hemicellulose fraction of lignocellulosic biomass (LCB) and of great interest for the green economy. Unfortunately, most of the cell factories cannot inherently metabolize xylose as sole carbon source. Yarrowia lipolytica is a non-conventional yeast that produces industrially important metabolites. The yeast is able to metabolize a large variety of substrates including both hydrophilic and hydrophobic carbon sources. However, Y. lipolytica lacks effective metabolic pathway for xylose uptake and only scarce information is available on utilization of xylose. For the economica feasibility of LCB-based biorefineries, effective utilization of both pentose and hexose sugars is obligatory.ResultsIn the present study, succinic acid (SA) production from xylose by Y. lipolytica was examined. To this end, Y. lipolytica PSA02004 strain was engineered by overexpressing pentose pathway cassette comprising xylose reductase (XR), xylitol dehydrogenase (XDH) and xylulose kinase (XK) gene. The recombinant strain exhibited a robust growth on xylose as sole carbon source and produced substantial amount of SA. The inhibition of cell growth and SA formation was observed above 60 g/L xylose concentration. The batch cultivation of the recombinant strain in a bioreactor resulted in a maximum biomass concentration of 7.3 g/L and SA titer of 11.2 g/L with the yield of 0.19 g/g. Similar results in terms of cell growth and SA production were obtained with xylose-rich hydrolysate derived from sugarcane bagasse. The fed-batch fermentation yielded biomass concentration of 11.8 g/L (OD600: 56.1) and SA titer of 22.3 g/L with a gradual decrease in pH below 4.0. Acetic acid was obtained as a main by-product in all the fermentations.ConclusionThe recombinant strain displayed potential for bioconversion of xylose to SA. Further, this study provided a new insight on conversion of lignocellulosic biomass into value-added products. To the best of our knowledge, this is the first study on SA production by Y. lipolytica using xylose as a sole carbon source.
Highlights
Introduction of xylose metabolic pathway inY. lipolytica PSA02004 As shown in previous section, Y. lipolytica PSA02004 strain showed no growth on minimal medium supplemented with 20 g/L xylose as a sole carbon source
Glucose and glycerol are the preferred carbon sources for Y. lipolytica, and these carbon sources were completely depleted within 72 h concomitant with the cell growth, which coincided with Succinic acid (SA) production (5.0-6.0 g/L) (Fig. 2a, c)
This was supported by similar cell growth (OD600: 20–22) observed on glucose/ glycerol, as well as during co-fermentation with xylose, where xylose was mainly utilized for xylitol synthesis and not contributing for biomass/product manufacturing
Summary
Introduction of xylose metabolic pathway inY. lipolytica PSA02004 As shown in previous section, Y. lipolytica PSA02004 strain showed no growth on minimal medium supplemented with 20 g/L xylose as a sole carbon source. The inability of Y. lipolytica to assimilate xylose for cellular growth impedes its application for lignocellulosic biorefineries. With the overexpression of XR, XDH and XK, the strain was able to grow in the medium containing xylose as a sole carbon source. The slightly high XDH activity than XR allows better synchronization between two enzymes, and results in efficient conversion xylose to xylulose without accumulation of xylitol as by-product. Y. lipolytica lacks effective metabolic pathway for xylose uptake and only scarce information is available on utilization of xylose. More attention is paid to the rewiring of metabolic networks of microbial strains to utilize multiple carbon sources simultaneously, especially glucose and xylose, from the feedstock which will be essential for derisking the commercial viability of the bioprocesses [2, 4]. It is envisaged that with the use of low-cost agricultural feedstock, bioproduction will soon replace the conventional petroleumbased process [9, 10]
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