Journal Article

Increased production of isobutanol from xylose through metabolic engineering of Saccharomyces cerevisiae overexpressing transcription factor Znf1 and exogenous genes

Soontorngun, Nitnipa, Songdech, Pattanan, Promdonkoy, Peerada, Butkinaree, Chutikarn, Yingchutrakul, Yodying, Runguphan, Weerawat

2024

Overview

Only trace amount of isobutanol is produced by the native Saccharomyces cerevisiae via degradation of amino acids. Despite several attempts using engineered yeast strains expressing exogenous genes, catabolite repression of glucose must be maintained together with high activity of downstream enzymes, involving iron–sulfur assimilation and isobutanol production. Here, we examined novel roles of nonfermentable carbon transcription factor Znf1 in isobutanol production during xylose utilization. RNA-seq analysis showed that Znf1 activates genes in valine biosynthesis, Ehrlich pathway and iron–sulfur assimilation while coupled deletion or downregulated expression of BUD21 further increased isobutanol biosynthesis from xylose. Overexpression of ZNF1 and xylose-reductase/dehydrogenase (XR-XDH) variants, a xylose-specific sugar transporter, xylulokinase, and enzymes of isobutanol pathway in the engineered S. cerevisiae pho13gre3Δ strain resulted in the superb ZNXISO strain, capable of producing high levels of isobutanol from xylose. The isobutanol titer of 14.809 ± 0.400 g/L was achieved, following addition of 0.05 g/L FeSO4.7H2O in 5 L bioreactor. It corresponded to 155.88 mg/g xylose consumed and + 264.75% improvement in isobutanol yield. This work highlights a new regulatory control of alternative carbon sources by Znf1 on various metabolic pathways. Importantly, we provide a foundational step toward more sustainable production of advanced biofuels from the second most abundant carbon source xylose. The engineered yeast strain could be used as a cell factory to produce high levels of isobutanol from the second most abundant sugar xylose due to activation of Znf1 target genes in the the valine biosynthesis, Ehrlich pathway and iron–sulfur assimilation.

• Publisher: Oxford University Press
• Subject: Biofuels / Bioreactors / Biosynthesis / Butanols / Carbon / Carbon - metabolism / Carbon sources / Catabolite repression / DNA-Binding Proteins - genetics / Enzymes / Fermentation / Gene silencing / Iron - metabolism / Metabolic Engineering / Metabolic pathways / Metabolism / Saccharomyces cerevisiae / Saccharomyces cerevisiae - metabolism / Saccharomyces cerevisiae Proteins - genetics / Saccharomyces cerevisiae Proteins - metabolism / Sulfur - metabolism / Transcription factors / Transcription Factors - genetics / Transcription Factors - metabolism / Valine / Xylose / Xylose - metabolism / Xylulokinase / Yeast