Integrated proteomics and metabolomics reveal how functional protein supplementation enhances salt tolerance in Zygosaccharomyces rouxii

Zygosaccharomyces rouxii is a halotolerant yeast widely associated with traditional high-salt food fermentations, while also being recognized as a spoilage microorganism in some food systems. Although its performance in fermented foods is strongly influenced by high salinity, the mechanisms by which nutritional inputs improve halotolerance remain unclear. Using DIA proteomics and UPLC-Orbitrap metabolomics, we evaluated Z. rouxii CGMCC 3791 subjected to 120 g/L NaCl with or without functional protein supplementation. Multi-omic remodelling was demonstrated by PCA, which showed distinct, repeatable separation of the control, salt, and supplemented groups. According to KEGG analysis, supplementation improved oxidative phosphorylation, ABC transporters, cofactor-related processes (one-carbon, vitamin B6, pantothenate-CoA), and amino acid pathways (arginine biosynthesis and arginine/proline metabolism), and preferentially enriched glycerolipid, glycerophospholipid, and GPI-anchor biosynthesis compared with salt alone. Coordinated changes in purine intermediates and energy carriers were revealed by integrated mapping, consistent with enhanced nucleotide homoeostasis and energy buffering. Correlation networks linked discriminating lipid-like metabolites to mitochondrial, redox, transport, and amino acid proteins. These findings indicate that functional protein supplementation improves halotolerance by combining membrane remodelling with metabolic and energetic reprogramming, offering options to optimise high-salt fermentations.