Simultaneous fermentation of galacturonic acid and five-carbon sugars by engineered Saccharomyces cerevisiae

•Engineered yeast consumes galacturonic acid, arabinose, and xylose simultaneously.•Six-carbon sugars inhibit galacturonic acid consumption.•Five-carbon sugars facilitate galacturonic acid consumption. Pectin-rich biomass has garnered attention as an alternative biomass source. However, some monomers derived from pectin-rich biomass, namely d-galacturonic acid, l-arabinose, and d-xylose, are not fermentable by industrial microorganisms such as Saccharomyces cerevisiae. The purpose of this study is to develop a S. cerevisiae strain capable of fermenting the pectin monomers. Expressions of eight heterologous genes and deletion of two endogenous genes, all of which were successfully completed by Cas9-based in vivo assembly and integration strategy, allowed the consumption of pectin monomers as sole carbon sources. To facilitate the consumption of galacturonic acid, which had the most limitations, the use of a co-substrate was tested using various sugars. As a result, we found that arabinose and xylose allowed simultaneous consumption of galacturonic acid. Based on intracellular metabolite profiling, it was concluded that the five-carbon sugars partially resolve the metabolic bottleneck of galacturonic acid.