Protein feed resources in China are scarce, which hinders the developement of feed industry and animal husbandry. Yeast protein is rich in nutrients and has significant development potential. Yeast strains were isolated and identified under acidic conditions (pH 3.5), from fermented flour, bovine rumen chyme, corn steep liquor, and other raw materials. Through the determination of the growth performance, bacterial protein and flavor of the strains, a promising high-protein yeast strain was identified as a potential strain for yeast protein processing. Further, the research indicated that 20 acid-tolerant yeast strains, including Saccharomyces cerevisiae, Pichia fermentans, Kluyveromyces marxianus, Yarrowia lipolytica, Candida tropicalis, Starmerella bacillaris, Zygosaccharomyces rouxii and Hanseniaspora thailandica, were obtained from different raw materials. Two superior strains, Pichia pastoris B-1 (7.65 g/L biomass, 4.96 g/L protein yield at pH 3.5; 9.32 g/L biomass, 5.81 g/L protein yield at pH 7.0) and Lactobacillus cerevisiae D-1 (8.00 g/L biomass, 3.91 g/L protein yield at pH 3.5; 14.91 g/L biomass, 6.87 g/L protein yield at pH 7.0), were identified based on biomass and protein production under shaking flask culture conditions. Flavor analysis revealed that strain B-1 had a higher content of flavor nucleotides and glutamic acid than strain D-1, along with a greater concentration of ester compounds. Thus, strain B-1 was identified as the potential strain for further application.. Therefore, B-1 strain was determined as the target strain. Finally, the growth characteristics and stress tolerance of B-1 strain were examined. It exhibited robust growth under conditions of pH≥2.0, temperature≤40 ℃ and NaCl concentration ( w /v )≤70 g/L.