Multi-omics-guided engineering of Saccharomyces cerevisiae to modulate higher alcohols and acetoin biosynthesis for improved wine sensory complexity

There is an urgent demand in the wine industry for effective strategies to reduce higher alcohol levels while enhancing flavor complexity without compromising sensory quality. Saccharomyces cerevisiae naturally produces acetoin, a desirable flavor compound, along with high levels of higher alcohols, which may cause dizziness and nausea. Here, we integrated multi-omics analysis with metabolic engineering to reprogram S. cerevisiae for enhanced production of acetoin and reduced formation of higher alcohols. Transcriptomic and metabolomic profiling under a gradient of inorganic nitrogen identified several differentially regulated pathways, including ribosome biogenesis, amino acid biosynthesis, carbon metabolism, and glycolysis. Based on pathway analysis, six key genes were selected as engineering targets: two NAD-dependent butanediol dehydrogenases (BDH1, BDH2), two transaminases (BAT1, BAT2), one alcohol dehydrogenase (ADH5), and one aldehyde dehydrogenase (ALD6). Eight recombinant strains (BY01-BY08) were constructed with different gene combinations. GC-MS analysis showed that strain BY07 achieved a 4.33-fold increase in acetoin, with reductions of 58.7 % in isobutanol, 23.2 % in active amyl alcohol, and 3.3 % in isoamyl alcohol, without affecting growth. This work not only advances our understanding of nitrogen-regulated acetoin and higher alcohols biosynthesis in S. cerevisiae but also provides an effective metabolic engineering strategy for enhancing flavor quality in industrial fermentations. • Adding NH 4 Cl modulates acetoin and higher-alcohols production in S. cerevisiae. • 350 mg/L NH 4 Cl gives max acetoin (1.13 g/L) and min higher alcohols (170.83 mg/L). • Multi-omics reveals 6 key genes ( BDH1/2 , BAT1/2 , ADH5 , ALD6 ) for pathway regulation. • Strain BY07 increases acetoin 4.33-fold and reduces isobutanol by 58.7 %.