Xylose Fermentation Was Improved by Kluyveromyces marxianus KHM89 through Up-regulation of Nicotinamide Adenine Dinucleotide (NAD⁺) Salvage Pathway

초록

Xylose fermentation has been reported to be improved in Kluyveromyces marxianus via strain improvement by overexpressing xylose reductase (XYL1) and xylitol dehydrogenase (XYL2). This study performed directed evolution to further enhance xylose consumption in a K. marxianus mutant following transcriptomic analysis to determine genes associated with enhanced characteristics. KmXYL1 and KmXYL2 genes were overexpressed in K. marxianus 17555 Delta URA3 for improving xylose fermentation. By performing directed evolution, a mutant K. marxianus KHM89 showing enhanced ethanol production was isolated from xylose medium. K. marxianus KHM89 consumed 47.39 g/L of xylose and produced 22.62 g/L of xylitol and 10.59 g/L of ethanol while the parental strain consumed 25.15 g/L of xylose and produced 7.36 g/L of xylitol and 2.05 g/L of ethanol. RNA sequencing-based transcriptomic analysis showed that alcohol dehydrogenases, aldehyde dehydrogenases, and NAD(+) salvage pathway enzymes were upregulated in K. marxianus KHM89. These results were achieved via a combinatorial approach of rational design and directed evolution. The findings of this study contribute to the improvement of xylose fermentation by K. marxianus at an industrial scale.

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