Proteomic Characterization of Microbiota-Derived Extracellular Vesicle-Enriched Fractions Detected in Red Yeast Rice

Fermented foods confer health benefits partially through microbial extracellular vesicles (EVs); however, complex fermented food-derived EVs remain understudied. In this research, we isolated and characterized microbiota-derived EV-enriched fractions (REVs) from 24 red yeast rice (RYR) samples obtained from three Chinese regions. The morphological analysis showed that the EV-enriched fractions exhibited a typical spherical, lipid bilayer structure. Proteomic profiling identified 2376 proteins, including 273 microbiota-derived proteins primarily originating from Monascus, Pectobacterium, and Erwinia species, with 143 proteins showing differential expression among regions. Functional analyses revealed significant enrichment in transport, glycolysis, translation, RNA degradation, and amino acid metabolism. Furthermore, we identified candidate region-associated proteins: rpsK and atpD in the FJ region; ypt-1, ALA1, and MIP11 in the GX region; and trxA, BAB1_1794, pstS, ugpB, tal, and glpK in the ZJ region. As the first comprehensive proteomic characterization of REVs, this study provides a basis for future studies on region-associated protein signatures and the potential application of fermented food-derived EVs in functional foods.